Western Blotting Brochure by KPL, Inc.

Western Blotting Some protein detection kits make you wonder if your eyes are playing tricks on you. SEE MORE with KPL!

Western Blotting No matter which KPL Western Blotting system you use, you’ll get a strong, clean signal every time. KPL’s blotting substrates and Protein Detector™ Western Blot Kits provide consistent, high quality results when analyzing proteins immobilized on membrane. Choose from a line of substrates and kits for either chemiluminescent or colorimetric methods of detecting specific antigens in a complex protein mixture. The cornerstone of KPL’s Western blotting systems is our line of sensitive liquid substrates. You have a choice among multiple chemiluminescent and chromogenic substrates

for peroxidase and phosphatase that provide varied levels of sensitivity. The chemiluminescent substrates provide a reliable alternative to conventional colorimetric ELISA with the advantage of increased sensitivity. Our chemiluminescent substrates for alkaline phosphatase detection, PhosphaGLO™ and PhosphaGLO Reserve™ AP Chemiluminescent Substrates, offer sensitivity and duration of signal in a one-component system.

KPL’s substrates are offered as standalone reagents or combined with unique blocking and wash solutions and highly specific secondary antibodies to create comprehensive, fully optimized kits for Western blotting. Each of these kits is designed to deliver the maximum signal-to-noise ratio available. Choose the detection system that best meets your needs and achieve clean, clear results blot after blot.

Substrates for Western Blotting KPL’s Western blotting substrates for use in peroxidase and phosphatase reporter systems offer high quality, reproducible chemiluminescent and colorimetric detection. It’s your choice! LumiGLO  HRP Reserve Substrate

LumiGLO HRP Substrate

PhosphaGLO  AP Reserve Substrate

 TMB HRP PhosphaGLO AP Substrate Substrate

4 CN Substrate

BCIP/NBT AP Substrate

 FirePhos Membrane AP Substrate

Type

chemi.

color.

Format

2-comp

1-comp

1- or 3comp

2-comp

1- or 3comp

1-comp

Detection Method

film or image analysis

visual-dark blue visual - purple precipitate precipitate

visual - purple precipitate

visual - red

Detection Limit

Sub-picogram; femtogram

0.6 picogram

femtogram

picogram

50 picogram

500 picogram

low nanogram

Stability of Working Solution

8 hours

24 hours

2 years at 4o C

1-comp1 year at 4o C. 3-comp24 hours

1 hour

1-comp1 year at 4o C. 3-comp1 hour

Duration of Signal

4 - 8 hours

1 - 2 hours

5 days

Enzyme Catalyst

HRP

Kinetics

5 minutes

15 minutes

10 minutes

Recommended Membrane

nitrocellulose or PVDF

comp = component • chemi. = chemiluminescent • color. = colorimetric • HRP = peroxidase • AP = phosphatase • NA = not applicable www.kpl.com

With KPLâ&#x20AC;&#x2122;s Protein Detectorâ&#x201E;˘ Western Blotting Systems, the results jump right out at you.

Western Blotting Chemiluminescent Substrates and Western Blot Kits Rapid, Sensitive Detection with Luminol-based Substrates Chemiluminescent detection involves the creation of light through the catalysis of an enzyme substrate. Use of this method for protein detection allows an increase in sensitivity by orders of magnitude compared to traditional colorimetric Western or dot blotting. KPL offers two unique luminol-based chemiluminescent substrates —LumiGLO® and LumiGLO Reserve™, for the rapid and sensitive detection of horseradish peroxidase (HRP)-labeled conjugates. Maximum Sensitivity with LumiGLO ReserveTM LumiGLO Reserve HRP Chemiluminescent Substrate offers an option for those assays where enhanced sensitivity is critical to success. This proprietary two-component substrate formulation provides greater than 20 times the sensitivity of standard LumiGLO with detection levels as low as the femtogram range (Figure 1).

Long Signal Duration LumiGLO Reserve emits light over the course of 4-8 hours with the most intense emission occurring within the first two hours. Because of its extreme light intensity, most images may be captured well under 10 minutes; multiple exposures are easy to obtain. High signal intensity facilitates the detection of both high and low abundance proteins and makes it an ideal system for use

Convenience LumiGLO Reserve Chemiluminescent Substrate Kits are supplied in several kit formats to meet diverse user needs. This substrate can replace your current chemiluminescent substrate in existing assays where greater sensitivity is desired. LumiGLO Reserve Western Blot Kit LumiGLO Reserve is also offered as part of a fully optimized kit, the Protein Detector LumiGLO Reserve Western Blot Kit, providing femtogram detection of proteins immobilized on membranes. For assurance against background interference, this kit contains our unique Detector™ Block for optimal signal-tonoise ratio; very low background can be achieved without compromise to signal intensity (Figures 2 and 3). Detector Block is also offered separately.

with chemiluminescent imagers. Sample and Antibody Conservation LumiGLO Reserve provides the added benefit of strong signal with the use of reduced amounts of precious target and antibodies. Therefore, material of limited supply or higher expense can be conserved while maintaining your current level of sensitivity. Superior Signal to Noise LumiGLO Reserve delivers lower nonspecific signal than competitor substrates in its class (Figures 2 and 3).

Reliable and Economical LumiGLO® KPL’s original LumiGLO Chemiluminescent Substrate provides high quality results in a variety of immuoassays.

LumiGLO Reserve Signal-to-Noise Comparison 1

LumiGLO B

LumiGLO Reserve C

ECL Plus

www.kpl.com

Figure 1: Relative expression of transcription factor, c-myc, using different chemiluminescent substrates. Five two-fold serial dilutions of purified c-myc (25 ng–1.56 ng, lanes 1–5) were compared to a 64 µg total protein HeLa nuclear lysate (lane 6). Following separation on a 4-20% PAGE gel and transfer to PVDF, protein was detected using a rabbit anti-c-myc antibody (1:200) and anti-rabbit HRP conjugate (1:10,000). Detection conditions were identical with the exception of substrate. While the c-myc lysate sample was not detectable with A) LumiGLO or C) ECL Plus™ after 10 minutes, the sample was easily detected with B) LumiGLO Reserve after just a 2-minute film exposure.

LumiGLO Reserve

ECL Plus

ECL Advance

Figure 2: Comparison of low-end sensitivity using LumiGLO Reserve and ECL Detection Kits. Two-fold serial dilutions of Mouse IgG (1 ng – 31 pg) were separated by SDS-PAGE and transferred to PVDF. Under manufacturer’s recommended conditions, protein was detected using HRP-labeled anti-mouse antibody (varied dilutions according to recommended optimization) and each respective substrate: A) LumiGLO Reserve, B) ECL Plus™, C) ECL Advance™. Film was exposed for 10 minutes and analyzed for sensitivity and signal to noise.

LumiGLO vs. Leading Traditional Chemiluminescent Substrates Superior Signal to Noise with Detector™ Block

Densitometry for LumiGLO vs. ECL 50000 LumiGLO

45000

ECL

Relative Image Density

40000 35000 30000 25000

LumiGLO Chemiluminescent Peroxidase Substrate

Supersignal® West Pico

ECL Plus™

ECL™

20000 15000 10000 5000 0 0

100

150

200

250

Concentration (ng)

Figure 4: Signal response comparison of LumiGLO vs. ECL™ chemiluminescent substrates in Western blotting. ß-galactosidase was electrophoresed, transferred to PVDF membrane and subsequently detected using rabbit anti-ß-gal followed by HRP conjugated goat anti-rabbit IgG (H+L). Each blot was treated with 5 mL of LumiGLO or ECL substrate and exposed to film for 10 minutes. The density of each band was analyzed on a Syngene GeneGenius™ image analyzer, using automatic background subtract.

Researchers continue to select LumiGLO for reliable performance in Western blot detection. Ideal for Routine Western Blotting

LumiGLO detects low picogram quantities of target protein on blots. After reaction with membrane-bound HRP conjugates, light emission begins immediately and continues for approximately 1–2 hours. Detection is possible within minutes of blot exposure.

Figure 5: Detection of ß-galactosidase in Western blots using alternative chemiluminescent substrates. A two-fold dilution series of purified ß-galactosidase (from 25 was electrophoresed on a polyacrylamide gel and transferred to KODAK BIOMAX Multi-Blot Kit for Proteins. Each blot was detected under the same conditions using Protein Detector LumiGLO Western Blotting Kit, substituting 5 mL LumiGLO on 3 of the 4 blots with 5 mL of leading competitive chemiluminescent substrates, respectively. Following a 10 minute film exposure, results were evaluated for sensitivity and signal:noise ratio.

Greater Linearity

LumiGLO produces a quantitatively linear signal on film that ensures a broader dynamic range of detection. While the light output from other chemiluminescent substrates tends to reduce sharply as the concentration of protein is titrated, a proportional reduction in sensitivity is achieved with LumiGLO (Figure 4). Low Background

LumiGLO delivers superior signal-tonoise by design. When used with KPL’s Detector™ Block, nonspecific binding is

further reduced and ensures low background without loss of signal intensity. Figure 5 compares the superior signalto-noise from blots blocked with Detector Block and subsequently detected with LumiGLO to leading competitor substrates. Cost Effective

LumiGLO is more economical blot for blot compared to competitive products. Enjoy the benefits of a sensitive, reliable substrate while staying within budget. LumiGLO Western Blot Kit

Figure 3: Signal linearity and signal-to-noise ratio comparison of advanced chemiluminescent substrates. Mouse IgG (250 pg to 3.91 pg) was transferred to nitrocellulose. Blots were detected under identical conditions with a 1:10,000 dilution of HRP-Goat-anti-Mouse IgG (except ECL Advance, 1:100,000) and exposed to film for 10 minutes. Densitometry was then performed using the Syngene GeneGenius. LumiGLO Reserve demonstrates superior signal linearity over a larger dynamic range and the greatest comparative signal-to-noise.

The LumiGLO Western Blot Kit is designed for low picogram detection of proteins immobilized on membranes. Researchers choose this kit for its reliable, consistent performance. The combination of a stable, liquid conjugate and a sensitive chemiluminescent substrate allows rapid and accurate identification of samples. For assurance against background interference, this kit contains our unique Detector Block for optimal signal-tonoise ratio (Figure 5).

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Western Blotting Sensitivity and Convenience with PhosphaGLOTM AP Substrates KPL offers two chemiluminescent substrates for use with alkaline phosphatase, PhosphaGLO ReserveTM and PhosphaGLOTM AP Substrates. PhosphaGLO Reserve Substrate overcomes the limitations posed by conventional chemiluminescent substrates for AP. With sensitivity in the femtogram range, PhosphaGLO Reserve enables detection of target protein in low concentrations. PhosphaGLO AP Substrate is recommended for routine detection of proteins in the picogram range (Figure 6). Low Background without Special Blockers

PhosphaGLO Substrates produce superior signal and low background, providing a better signal to noise ratio and a cleaner image than other chemiluminescent substrates for AP (Figure 6). No special blockers are needed with either nitrocellulose or PVDF membrane. Ultimate Convenience

Both PhosphaGLO AP Substrates offer exceptional convenience as one-component solutions ready to use. They are stable for up to two years when stored at 4o C. Their long glow times of 5 days facilitate assay development, enabling repeat exposures.

PhosphaGLO Reserve AP Substrate

PhosphaGLO AP Substrate

Colorimetric Kits

Western Blot Reagents

KPL also offers colorimetric Western blot systems when chemiluminescent detection is not preferred. Results can be interpreted by direct visualization of the blot. Everything you need to detect your antigen and primary antibody is supplied for convenience and optimal performance.

In addition to our line of substrates and kits, KPL offers a broad line of secondary antibodies, conjugates and support reagents in various packaging formats to suit your needs.

Protein DetectorTM TMB Western Blot Kit The Protein DetectorTM TMB Western Blot Kit utilizes TMB Peroxidase Membrane Substrate (3,3’,5,5’-tetramethylbenzidine), the most sensitive chromogenic peroxidase substrate for Western and dot blotting applications. Detection limits are significantly increased as compared to other chromogenic membrane substrates. TMB produces a dark blue precipitate upon reaction with HRP.

Secondary Antibodies

• Highly purified and specific. • Large offering of species-specific polyclonal antibodies • Enzyme, biotin and fluorochrome-labeled antibodies Support Reagents

• One source for block solutions, wash buffers, diluents and stabilizers. • Consistently produced for reliable results.

Protein DetectorTM BCIP/NBT Western Blot Kit For detection of phosphatase-labeled conjugates, the Protein Detector BCIP/NBT Western Blot Kit is ideal. When reacted with alkaline phosphatase, BCIP/NBT produces clean, intense bands of purple precipitate. It also provides stable, more permanent results than other chromogenic substrates.

CDP Star® Chemiluminescent Substrate

Figure 6: Comparison of low-end sensitivity using PhosphaGLO Reserve AP Substrate, PhosphaGLO AP Substrate, and CDP-Star Chemiluminescent Substrate on PVDF membranes. Five-fold serial dilutions of mouse IgG (2 ng -3.2 pg) were separated by SDS-PAGE and transferred to membranes. Protein was detected using a 1:1000 dilution of biotin-labeled goat anti-mouse IgG, a 1:10,000 dilution of AP-labeled streptavidin and each respective substrate. Film was exposed for 10 minutes.

LumiGLO is a registered trademark and LumiGLO Reserve, Protein Detector, PhosphaGLO, PhosphaGLO Reserve, FirePhos and Detector are trademarks of KPL, Inc. ECL, ECL Plus and ECL Advance are trademarks of GE Healthcare. SuperSignal West is a registered trademark of Pierce Biotechnology, Inc. BIOMAX is a trademark of KODAK. GeneGenius is a trademark of Syngene. Biodyne is a registered trademark of Pall/Gelman Corporation. CDP-Star is a registered trademark of Applied Biosystems.

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AP Membrane Color Substrates See RED and PURPLE in your Western blots! BCIP/NBT (5-Bromo-4-chloro-3-indolyl phosphate/ nitroblue tetrazolium) is the most commonly used substrate for alkaline phosphatase (AP) detection in Western blotting due to its high sensitivity and low background staining. Traditional formulations of BCIP/NBT produce a deep purple color when reacted with AP conjugates. Now KPL’s new BCIP/INT formulation, FirePhosTM AP Membrane Substrate, forms a red precipitate, providing a choice of red or purple precipitating AP substrates to better optimize your individual assays. FirePhosTM AP Membrane Substrate BCIP/NBT Phosphatase Substrate Sensitive and reproducible

(Figure 1) , sharp band resolution and a clear image. The color produced is stable and linear over a wide dynamic range.

FirePhos AP Membrane Substrate

BCIP/NBT Substrate (1-Component)

BCIP/NBT Substrate (3-Component)

Figure 1: KPL’s substrates for alkaline phosphatase detection in Western blotting demonstrate high sensitivity and low background. A two-fold dilution series from 250 ng to 3.9 ng of mouse IgG was loaded onto a Tris-HCl gradient gel, then blotted onto nitrocellulose. The antigen was detected with ReserveAPTM Goat Anti-mouse Conjugate (KPL) followed by FirePhos AP Membrane and BCIP/NBT Substrates (1component and 3-component).

No matter which substrate you choose, you’ll achieve sensitive and reproducible detection of proteins on Western blots. Both substrates provide sensitivity in the low nanogram range with minimal background SEE MORE with KPL!

ReserveAP Conjugates

AP Membrane Color Substrates Non-fading

Improve your assay performance even further with ReserveAPTM-labeled Antibodies!

FirePhos and BCIP/NBT produce a bright red or purple precipitate that can be deposited on nitrocellulose and PVDF membranes. They resist fading when exposed to light. Convenient

KPL’s new ReserveAPTM Alkaline

These substrates are supplied as convenient one-component solutions that are ready to use. KPL’s BCIP/NBT is also available as a three-component solution. Both substrates are provided in several sizes to meet your needs.

Phosphatase-labeled Conjugates exhibit high potency and consistent performance in immunoassays. These conjugates are the result of advances in our conjugation chem-

Ordering Information Catalog#

istry and offer high signal and low

Description

Size

background while meeting KPL’s standards for stability and repro-

FirePhosTM AP Membrane Substrate 50-81-30

FirePhos Membrane Phosphatase Substrate (1-Component)

100 mL

ducibility. They are ideal for

50-81-40

FirePhos Membrane Phosphatase Substrate (1-Component)

400 mL

demanding immunoassays that

50-81-34

FirePhos Membrane Phosphatase Substratee (1-Component)

require high detection sensitivity, including ELISA, Western blotting

BCIP/NBT Phosphatase Membrane Substrate

and immunohistology. Visit

50-81-18

BCIP/NBT Membrane Phosphatase Substrate (1-Component)

100 mL

www.kpl.com for more information

50-81-07

BCIP/NBT Membrane Phosphatase Substrate (1-Component)

600 mL

on KPL’s ReserveAP conjugates.

50-81-10

BCIP/NBT Membrane Phosphatase Substrate (1-Component)

50-81-00

BCIP/NBT Membrane Phosphatase Substrate (3-Component)

300 mL

Detector Block (5X)

240 mL

Membrane Blocking 71-83-00

KPL’s colorimetric membrane substrates are part of our comprehensive line of Western blottng kits and reagents. Ask about our AP chemiluminescent substrates that can significantly lower the detection limit of an assay. SEE MORE with KPL!

Gaithersburg, MD 20878 Phone: 800.638.3167 Fax: 301.948.0169 www.kpl.com

To order or for more information, contact us at 800.638.3167 / 301.948.7755, fax 301.948.0169 or visit us at www.kpl.com.

ISO 9001:2000 Registered

HRP Membrane Color Substrates Sensitivity, reliability and affordability! KPL offers two substrates commonly used to detect

price. The substrate may be adapted as a soluble sub-

horseradish peroxidase (HRP) conjugated antibodies

strate for ELISA by omitting TMB Membrane

that bring complementary benefits to membrane

Enhancer. Both substrates perform equally well on

detection; TMB (3, 3´, 5, 5´-tetramethylbenzidene)

nitrocellulose and PVDF membranes.

Membrane Substrate and 4 CN (4-chloro-1-naphthol) Membrane Substrate. TMB substrate offers the most

4 CN produces a distinct, blue-purple precipitate and extremely low background.

sensitive colorimetric detection for blotting applications and is recommended when high signal and low background are required. 4 CN substrate is highly reliable and is notable for its characteristically low

TMB produces a blue precipitate and provides greater sensitivity than 4 CN.

background. •

TMB Membrane Substrates (1- and (3-Component) 4 CN

TMB

In the presence of peroxidase conjugate, KPL’s TMB Membrane Substrates produce a stable, dark blue pre-

• 4 CN Peroxidase Substrate (2-Component)

cipitate at the site of a positive reaction, enabling detection of as little as 50 pg of peroxidase. Our TMB

4 CN (4-chloro-1-napthol) Peroxidase Membrane

substrates are available as 1- and 3- component solu-

Substrate produces a purple precipitate at the reaction

tions. 1-component TMB Peroxidase Substrate is ready

site in the presence of peroxidase conjugates. Due to

to use and contains hydrogen peroxide and buffer.

its low background, 4 CN presents a desirable alter-

Our 3-component TMB contains TMB in an organic

native to other, more sensitive substrates when back-

base, H2O2 in a citric acid buffer and a third component, TMB Membrane Enhancer. It provides sensitivi-

ground poses a problem.

ty equivalent to 1-component TMB at an economic

SEE MORE with KPL!

Support Reagents Benefits of KPL’s HRP Membrane Substrates •

KPL provides a broad range of assay support reagents for Western blot detection. Conjugate stabilizers, diluents, block and wash solutions are offered in convenient stable liquid form to ensure accurate reproducible results, blot after blot. Our Detector Block is ideal for use as a diluent/block solution for Western blots, Southern blots and Northern blots on a variety of membranes.

TMB Peroxidase Membrane Substrate

• • • • • • • •

Most sensitive colorimetric substrate for blotting Delivers results at the picogram level Recommended for both kinetic and endpoint ELISA Contains no harmful organic solvents; safer and more sensitive than DAB or AEC. 1-Component TMB-no preparation required. Ensures more consistent results 3-Component TMB-same performance as 1-Component TMB at an economic price. Blots remain stable with minimal fading

4 CN Peroxidase Substrate

• • •

• DetectorTM Block

Easy mix-and-use, two-component solutions Very low background, moderate sensitivity High contrast image ideal for photography and publication

Ordering Information Catalog#

Description

Size

50-77-18

TMB Membrane Substrate (1-Component)

100 mL

50-77-03

TMB Membrane Substrate (1-Component)

200 mL

50-77-04

TMB Membrane Substrate (1-Component)

50-77-00

TMB Membrane Substrate (3-Component)

440 mL

50-77-01

TMB Membrane Enhancer

50-73-00

4 CN Substrate System (2-Component)

600 mL

50-73-04

4 CN Substrate System (2-Component)

2700 mL

Detector Block is recommended as a general blocking agent in membrane-based detection assays. It is especially useful when traditional blocking solutions (i.e., milk, BSA, casein) reduce sensitivity or do not adequately block background. Detector Block can be used as both a blocking solution and a conjugate diluent in membrane applications.

40 mL

Protein Detector TMB Western Blot Kit 54-11-50

Protein Detector TMB Western Blot Kit

2500 cm2

Detector Block 71-83-00

Detector Block

240 mL

Gaithersburg, MD 20878 Phone: 800.638.3167 Fax: 301.948.0169

To order or for more information on KPL’s line of protein detection products, contact us at 800.638.3167 / 301.948.7755, fax 301.948.0169 or visit us at www.kpl.com.

www.kpl.com

ISO 9001:2000 Registered

Immunoassay Buffers Save preparation time with KPL’s new line of quality buffers! Lighten your work load with KPL’s new line of quality

registered quality management system. All buffers are con-

buffers. Offered as convenient liquid concentrates, they

ductivity controlled. KPL offers a variety of buffers designed

eliminate the need for time-consuming buffer preparation.

for use in general immunoassay applications such as Western

KPL buffers are manufactured and carefully controlled for

blotting, gel electrophoresis, ELISA and sample preparation.

quality and consistent performance with our ISO 9001:2008Description

1X Composition

Applications

10X Tris-Glycine Transfer Buffer

25 mM Tris, 192 mM glycine. pH 8.3

10X Tris-Glycine-SDS

25 mM Tris, 192 mM glycine, 0.1% SDS. pH 8.3

For preparing a standard Western blot transfer buffer (Towbin) and as a gel electrophoresis buffer for native Tris-glycine gels without SDS. . Running buffer for sodium dodecyl sulfate – polyacrylamide gel electrophoresis (SDS-PAGE) of proteins.

10X Phosphate-Buffered Saline (PBS)

10 mM Phosphate,150 mM NaCl. pH 7.4

Phosphate-Buffered Tween 20, (PBST)

10 mM Phosphate, 150 mM NaCl, 0.05% Tween 20. pH 7.4

10X Tris-Buffered Saline (TBS)

50 mM Tris, 150 mM NaCl. pH 7.6

10% Tween 20

Wash buffer for general immunoassay applications and sample preparation. Where applicable, Tween 20 is a nonionic detergent that reduces nonspecific binding and protein-protein interaction during wash steps.

Nonionic detergent additive for PBS and Western blotting wash solutions. Reduces nonspecific binding and proteinprotein interactions.

See reverse side.

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Immunoassay Buffers

KPL Quality Buffers (cont’d) Description

1X Composition

Applications

5% SDS

5% Sodium Dodecyl Sulfate

Detergent surfactant used in preparing proteins for sodium dodecyl sulfate – polyacrylamide gel electrophoresis (SDS-PAGE) and as an additive to transfer buffers in Western blotting. SDS increases the elution rate of proteins from the gel.

20X SSC

15 mM sodium citrate, 150 mM NaCl in DEPC water. pH 7.0

Used in a variety of molecular biology applications. Facilitates transfer of nucleic acids to membranes.

10X Dulbecco's PBS (D-PBS)

8.5 mM sodium phosphate, 1.5 mM potassium phosphate, 137 mM NaCl. pH 7.4

For sample preparation and as a wash buffer in general immunoassay, tissue, and cell culture applications. Not formulated with magnesium or calcium salts. Where applicable, Tween 20 is a non-ionic detergent that reduces nonspecific binding.

Other Immunoassay Support Reagents Available from KPL ELISA and Western Blotting Applications: • AP and HRP Conjugate Stabilizers

10X Dulbecco's with Tween 20. (D-PBST)

8.5 mM sodium phosphate, 1.5 mM potassium phosphate, 137 mM NaCl, 2.7 mM KCl. 0.05% Tween 20. pH 7.4

10X Tris-Buffered Saline with 0.5% Tween 20 (TBST)

50 mM Tris, 150 mM NaCl, 0.05% Tween 20. pH 7.6

10X Tris-Buffered Saline with 10% Tween 20 (TBST)

50 mM Tris, 150 mM NaCl, 1.0% Tween 20. pH 7.6

• 10% BSA Diluent/Blocking Solution Concentrate • Milk/Diluent Blocking Solution Concentrate • 20X Wash Solution Concentrate

As a general wash buffer in immunoassay applications. Tween 20 is a non-ionic detergent that reduces nonspecific binding. The 10% Tween 20 formulation provides a more stringent wash than standard TBST formulations.

Western Blotting Applications: • Detector Block • Biotin Wash Kit

Take the time and effort out of your buffer preparation. Choose from our line of popular buffers and enjoy the benefits - convenience, reliability and confidence - that KPL’s pretested, quality buffers bring to your research.

ELISA Applications: • Coating Solution Concentrate • Stop Solutions

Ordering Information Catalog#

Description

Size

51-10-01 51-10-02 51-11-01 51-11-02 51-13-01 51-13-02 51-14-01 51-14-02 51-17-01 51-17-02 51-12-01 51-12-02 51-20-01 50-86-05 51-15-01 51-15-02 51-16-01 51-16-02 51-18-01 51-18-02 51-19-01 51-19-02

10X Tris-Glycine Transfer Buffer

1L 5L 1L 5L 1L 5L 200 mL 1L 1L 5L 200 mL 1L 200 mL 1L 1L 5L 200 mL 1L 200 mL 1L 200 mL 1L

10X Tris-Glycine SDS Buffer 10X Phosphate-Buffered Saline (PBS) 10X Phosphate-Buffered Saline with Tween 20 (PBST) 10X Tris-Buffered Saline (TBS) 10% Tween 20 5% SDS 20X SSC 10X Dulbecco's PBS (D-PBS) 10X Dulbecco's PBS with Tween 20 (D-PBST) 10X Tris Buffered Saline with 0.5% Tween 20 (TBST) 10X Tris Buffered Saline with 10% Tween 20 (TBST)

Gaithersburg, MD 20878 Phone: 800.638.3167 Fax: 301.948.0169 www.kpl.com

ISO 9001:2008 Registered

To order or for more information on KPL’s full line of protein detection products, contact us at 800.638.3167 / 301.948.7755, fax 301.948.0169 or visit us at www.kpl.com. ©2009 KPL, Inc. All rights reserved. ML363-03

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ReserveAP Conjugates TM

Spice up your assay with our red hot high potency ReserveAP Conjugates! KPLâ&#x20AC;&#x2122;s new ReserveAPTM Alkaline Phosphatase

line of AP conjugates in ELISA and outperform

(AP)-labeled antibody conjugates exhibit high

AP conjugates offered by other manufacturers.

potency and consistent performance in

Higher conjugate dilutability is also observed

immunoassays. These conjugates are the result

without loss of linearity, enabling precious

of advances in our conjugation technology and

antigen or primary antibody conservation.

offer higher signal than our current line of AP

Consistent Performance

conjugates while meeting the same standards for low background, stability and reproducibili-

Reproducible antibody conjugation and consis-

ty. They are intended for demanding

tent performance are verified according to our

immunoassays that require high detection sen-

ISO 9001:2000-registered quality assurance

sitivity, including ELISA, Western blotting and

system. Lot consistency studies in which three

immunohistology.

lots were studied by ELISA indicated minimal variability.

Higher Potency Excellent Value ReserveAP Conjugates are affinity purified and conjugated to the highest grade of alkaline

ReserveAP Conjugates provide high perform-

phosphatase. In our studies, they generate two-

ance at an economical price.

to-three fold higher values than our current SEE MORE with KPL!

AP Substrates Chemiluminescent ELISA: A constant amount of antigen (mouse IgG) was coated onto a microtiter plate. The plate was probed with varying concentrations of goat anti-mouse IgG (H+L) AP conjugate, including KPL’s standard AP conjugate and ReserveAP™ conjugate, followed by KPL’s PhosphaGLO™ Chemiluminescent AP Substrate.

KPL offers a range of sensitive substrates for the detection and quantification of phosphatase (AP) activity. They provide a choice of intense colors for ELISA and blotting applications.

ELISA •

Conclusion: The results indicate that ReserveAP™ conjugate offers superior performance to our standard AP conjugate with a ~3-fold lower amount of conjugate required for detection.

ReserveAPTM Conjugates Ordering Information

•

Catalog#

Description

Size

0751-1001

Goat Anti-Human IgA (α)

1.0 mg

0751-1002

Goat Anti-Human IgG (γ)

1.0 mg

0751-1003 0751-1004

Goat Anti-Human IgM (µ) Goat Anti-Human IgE (ε)

0751-1006

FirePhosTM AP Microwell Substrate BluePhos® AP Microwell Substrate pNPP Phosphatase Substrate

Blotting •

1.0 mg 1.0 mg

FirePhos AP Membrane Substrate

•

BCIP/NBT Phosphatase Substrate

Goat Anti-Human IgG (H+L)

1.0 mg

•

PhosphaGLO AP Substrate

0751-1007

Goat Anti-Human IgA+IgG+IgM (H+L)

1.0 mg

•

2151-1002

0.5 mg

4751-1002

F(ab’)2 Anti-Human IgG (γ) Goat Anti-Human IgG (γ) liquid

PhosphaGLOTM Reserve AP Substrate

4751-1003

Goat Anti-Human IgM (µ) liquid

1.0 mg

4751-1006

Goat Anti-Human IgG (H+L) liquid

1.0 mg

0751-1802

Goat Anti-Mouse IgG (γ) HSA

1.0 mg

0751-1803

Goat Anti-Mouse IgM (µ) HSA

1.0 mg

0751-1806

Goat Anti-Mouse IgG (H+L) HSA

1.0 mg

0751-1809

Goat Anti-Mouse IgG +IgM (H+L) HSA

1.0 mg

4751-1802

Goat Anti-Mouse IgG (γ) HSA, liquid

1.0 mg

4751-1806

Goat Anti-Mouse IgG (H+L) HSA, liquid

1.0 mg

151-18-01

Goat Anti-Mouse IgA (α) HSA

0.5 mg

0751-1807

Goat Anti-Mouse IgA+IgG+IgM (H+L) HSA

1.0 mg

0751-1506

Goat Anti-Rabbit IgG (H+L)

1.0 mg

0751-1516

Goat Anti-Rabbit IgG (H+L) HSA

1.0 mg

4751-1506

Goat Anti-Rabbit IgG (H+L), liquid

1.0 mg

4751-1516

Goat Anti-Rabbit IgG (H+L) HSA, liquid

1.0 mg

Whichever substrate you choose, enjoy the benefits of excellent signalto-noise with higher sensitivity and lower background than that of other AP substrates. Visit www.kpl.com for more information.

Gaithersburg, MD 20878 Phone: 800.638.3167 Fax: 301.948.0169 www.kpl.com ISO 9001:2000 Registered

HSA=Human Serum Adsorbed

Visit our website at www.kpl.com for a complete listing of ReserveAP conjugates. To order or for more information on KPL’s protein research products, contact us at 800.638.3167 / 301.948.7755, FAX 301.948.0169 or visit us at www.kpl.com. ML349-04 For research use only. ©2007 KPL, Inc. All rights reserved.

SEE MORE with KPL!

DyLightTM Fluorescent Conjugates KPL’s DyLightTM Conjugates - A Brilliant Choice! KPL’s DyLightTM conjugates offer a brilliant choice in a variety of multicolor detection applications, including fluorescence microscopy, flow cytometry, Western blotting, ELISA and array platforms. Our affinity purified antibodies combined with a series of outstanding DyLight dyes provide superior performance over conventional CyDyeTM fluors, fluorescein and rhodamine, with performance comparable to that of Alexa Fluor® dyes (Figure 1). Enjoy these advantages when you switch to KPL’s DyLight Conjugates:

KPL offers eight DyLight dyes, including 405, 488, 549, 594, 633, 649, 680 and 800 with well-differentiated excitation and emission spectra. Our extensive line of over 170 DyLight conjugates is available across a range of animal species immunoglobulin, including human, mouse, rabbit, rat, other species and streptavidin. See back cover to find out what sets KPL’s antibodies apart. Comparison of DyLight 649 and Alexa 647 Conjugates in a Performance Assay

• High sensitivity, bright signal enable conjugate conservation

• More photostable than Cy, FITC; comparable to Alexa High signal-to-noise ratios; low background Wide selection of specific antibodies Consistent, lot-to-lot performance Narrow emission spectra enable specific, multicolor analysis • Compatible with a variety of buffers.

• • • •

Antigen Concentration (ng/mL)

Figure 1. KPL DyLight 649 conjugates demonstrate comparable fluorescense intensity and photostability to Alexa 647 conjugates in a FLISA.

Image above: HMVEC-L primary endothelial cells. F-actin detected with DY554-Phalloidin (rendered green). Microtubules detected with anti-tubulin antibody and DyLight 649 conjugated goat anti-mouse IgG (red). Nuclei detected with DAPI (blue)

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DyLightTM Fluorescent Conjugates DyLight Conjugated Affinity Purified Antibodies and Streptavidin KPL DyLight conjugates provide a choice of outstanding fluorescent conjugates with absorption spectra ranging from 405 nm to 800 nm. Their emission profiles correspond to those of other commonly used fluorophores such as Alexa, FITC and the Cy dyes. Narrow, defined emission spectra enable the use of multiple fluorescence labeling and simultaneous identification of several target molecules in the same sample. Light output is comparable to IRDye and Alexa and more intense than Cy Dyes, FITC or TRITC. DyLight dyes demonstrate resistance to photobleaching, resulting in excellent photostability as well as high solubility in aqueous solutions across a range of pH values.

Emission

DyLight Dye

Ex/Em(nm) Replaces

Blue

DyLight 405

400/420

Alexa 405 and Cascade Blue

DyLight 488

493/518

Alexa 488, Fluorescein and FITC

DyLight 549

550/568

Alexa 546, Alexa 555, Cy3, TRITC

Green Yellow Orange

DyLight 594

593/618

Alexa 594, Texas Red

Red

DyLight 633

638/658

Alexa 633

Red

DyLight 649

646/674

Alexa 647, Cy5

Near IR

DyLight 680

682/715

Alexa 680, Cy5.5

Infrared

DyLight 800

770/794

IRDye 800

nm = nanometers IR = infrared

Ordering Information Description Anti-Mouse IgG (γ), HSA Anti-Mouse IgG (H+L), HSA Anti-Mouse IgG (H+L), HSA, 0.1 mg Anti-Mouse IgG (H+L), RbSA, HSA Anti-Mouse IgM (µ), HSA Anti-Mouse IgG+IgM (H+L), HSA F(ab’)2 Anti-Mouse IgG (γ), HSA F(ab’)2 Anti-Mouse IgG (H+L), HSA Anti-Human IgG (H+L) Anti-Human IgG (H+L), 0.1 mg Anti-Human IgG (γ) Anti-Human IgM (µ) F(ab’)2 Anti-Human IgG (γ) F(ab’)2 Anti-Human IgM (µ) F(ab’)2 Anti-Human IgG (H+L) Anti-Rabbit IgG (H+L) Anti-Rabbit IgG (H+L), 0.1 mg Anti-Rabbit IgG (H+L), HSA F(ab’)2 Anti-Rabbit IgG (H+L), HSA Anti-Rat IgG (H+L) Anti-Rat IgG (H+L), 0.1 mg Anti-Guinea Pig IgG (H+L) Anti-Chicken IgG (H+L) Anti-Dog IgG (H+L) Anti-Goat IgG (H+L) Anti-Horse IgG (H+L) Rabbit Anti-Sheep IgG (H+L) Anti-Swine IgG (H+L) Streptavidin Streptavidin, 0.1 mg

SEE MORE with KPL!

DyLight 405* DyLight 488 DyLight 549

072-08-18-06 042-08-18-06

202-08-18-06 072-08-10-06 042-08-10-06

202-08-10-06 072-08-15-06 042-08-15-06 202-08-15-16 072-08-16-06

072-08-24-06 072-08-13-06

072-08-30-00 042-08-30-00

*Coming soon!

072-03-18-02 072-03-18-06 042-03-18-06 072-03-18-18 072-03-18-03 072-03-18-09 202-03-18-02 202-03-18-06 072-03-10-06 042-03-10-06 072-03-10-02 072-03-10-03 202-03-10-02 202-03-10-03 202-03-10-06 072-03-15-06 042-03-15-06 072-03-15-16 202-03-15-16 072-03-16-06 042-03-16-06 072-03-17-06 072-03-24-06 072-03-19-06 072-03-13-06 072-03-21-06 072-03-23-06 072-03-14-06 072-03-30-00 042-03-30-00

DyLight 594*

DyLight 633* DyLight 649

072-04-18-02 072-04-18-06 042-04-18-06 072-04-18-18 072-04-18-03 072-04-18-09 202-04-18-02 202-04-18-06 072-04-10-06 042-04-10-06 072-04-10-02 072-04-10-03 202-04-10-02 202-04-10-03 202-04-10-06 072-04-15-06 042-04-15-06 072-04-15-16 202-04-15-16 072-04-16-06

072-05-18-02 072-09-18-06 072-10-18-06 042-09-18-06 042-10-18-06 072-05-18-18 072-05-18-03 072-05-18-09 202-05-18-02 202-09-18-06 202-10-18-06 072-09-10-06 072-10-10-06 042-09-10-06 042-10-10-06 072-05-10-02 072-05-10-03 202-05-10-02 202-05-10-03 202-09-10-06 202-10-10-06 072-09-15-06 072-10-15-06 042-09-15-06 042-10-15-06 072-05-15-16 202-09-15-16 202-10-15-16 072-09-16-06 072-10-16-06

072-04-17-06 072-04-24-06 072-04-19-06 072-04-13-06 072-04-21-06 072-04-23-06 072-04-14-06 072-04-30-00 042-04-30-00

072-05-17-06 072-09-24-06 072-10-24-06 072-05-19-06 072-09-13-06 072-10-13-06 072-05-21-06 072-05-23-06 072-05-14-06 072-09-30-00 072-10-30-00 042-09-30-00 042-10-30-00

DyLight 680

DyLight 800

072-05-18-06 042-05-18-06 072-06-18-18

072-06-18-06 042-06-18-06 072-07-18-18

072-07-18-06 042-07-18-06

202-05-18-06 072-05-10-06 042-05-10-06

202-06-18-06 072-06-10-06 042-06-10-06

202-07-18-06 072-07-10-06 042-07-10-06

202-05-10-06 072-05-15-06 042-05-15-06 072-06-15-16 202-05-15-16 072-05-16-06

202-06-10-06 072-06-15-06 042-06-15-06 072-07-15-16 202-06-15-16 072-06-16-06

202-07-10-06 072-07-15-06 042-07-15-06

072-06-17-06 072-05-24-06 072-06-19-06 072-05-13-06 072-06-21-06 072-06-23-06 072-06-14-06 072-05-30-00 042-05-30-00

072-07-17-06 072-06-24-06 072-07-19-06 072-06-13-06 072-07-21-06 072-07-23-06 072-07-14-06 072-06-30-00 042-06-30-00

202-07-15-16 072-07-16-06

072-07-24-06 072-07-13-06

072-07-30-00 042-07-30-00

Near Infrared and Infrared Fluorophores DyLight 680 and 800 Conjugates Offer Sensitive Multicolor Imaging in Western Blotting

DyLight Dyes Emission Spectra

DyLight 680 and 800 dyes emit in the near infrared and infrared ranges of the light spectrum respectively and are ideal for multicolor protein detection in Western blotting. Unlike fluorescent conjugates that emit in the visible range, DyLight 680 and 800 conjugates provide a unique set of advantages: • Secondary antibodies with defined specificity and

sensitivity • Brighter signal than visible fluorescence • Virtually no background autofluorescence from membranes or most biological specimens Effective Alternative to Chemiluminescence • Broader dynamic range than chemiluminescence • Quantitation accuracy superior to traditional

methods • Easy to assay multiple proteins simultaneously on one Western blot • Cost-effective - eliminates need for chemilumin-

escence substrates, film and darkroom • Clean results - no bleeding from consecutive lanes

Near Infrared Fluorescent Imaging with Secondary Antibodies labeled with DyLight 680 and 800

Figure 2. DyLight 680-anti mouse IgG and DyLight 800 anti-rabbit IgG secondary antibody conjugates provide low background and high signal in two-color Western blot detection of tubulin and TNFα. Membrane was imaged with the LI-COR Odyssey Infrared Imaging System. HSA = human serum adsorbed RbSA = rabbit serum adsorbed DyLight antibody conjugates are made in goat except anti-goat and anti-sheep antibodies are made in rabbit. Supplied in 1.0 mg lyophilized form except select 0.1 mg sizes.

1.800.638.3167

www.kpl.com

Immunofluorescence using DyLight Conjugates See bright fluorescence and low background with KPL DyLight conjugates in immunohistology applications. The winning combination of DyLight dyes and KPL purified antibodies enables multicolor labeling of two or more targets with similar intensity and photostability to Alexa dyes without the limitations of fluorescein and CyDyes.

Mouse primary cortical neurons MCx WCS stained with DyLight 549 (red). Synaptophysin stained with DyLight 488 (green).

HMVEC-L primary endothelial cells F-actin detected with DY554-Phalloidin (rendered green). Microtubules stained with DyLight 649 (red). Nuclei detected with DAPI (blue).

Rat hippocampal neurons WCS stained with; DyLight 488 (green). MAP2 stained with DyLight 549 (red).

A549 cells Cytokeratin stained with DyLight 680 (pseudocolored white). Lamin A stained with DyLight 549 (pseudocolored red).

DyLightTM Fluorescent Conjugates KPL AntibodiesWhat Sets Them Apart In 1979, KPL pioneered the production of large-scale affinity purification and was the first company to commercialize affinity purified secondary antibodies. Rigorous standards throughout the antibody production process make our antibodies standout in the marketplace. Many manufacturers cut corners by beginning with inferior serum and extracting the useful antibody towards the end of their process. KPL spends considerable effort developing and purifying its own immunogen formulation to generate the antiserum, because pure immunogen results in a more potent and specific antibody prior to any purification steps. Further, ISO 9001:2008-certified quality procedures are carried out at more than six different stages of the antibody production cycle, and material that does not meet our high standards for potency and cross-reactivity is rejected. Our late stage purification process has been continually refined since 1979 and relies on a one of a kind custom column resin which is uniquely suited to our antibody manufacturing process. Finally, our process of pooling antiserum from multiple animals tempers natural serum variability, minimizing variances from animal-to-animal. The result is more standardized large-scale antibody lots with increased consistency. KPL’s experience, innovative processes, and attention to detail result in highperformance antibodies that are unique.

NIH 3T3 cells F-actin detected with DY554-Phalloidin (rendered green). Microtubules stained with DyLight 649 (red). Nuclei detected with DAPI (blue) Gaithersburg, MD Phone: 800.638.3167/301.948.7755 Fax: 301.948.0169 www.kpl.com

To order or for more information on KPL’s line of unlabeled and conjugated affinity purified antibodies, contact us at 800.638.3167/301.948.7755,

ISO 9001:2008 Registered

fax 301.948.0169 or visit www.kpl.com. ML356-02 For research use only. DyLight is a trademark of Thermo Fisher Scientific Inc. and its subsidiaries. Cy and Cy Dye are trademarks of GE Healthcare. Alexa Fluor is a registered trademark of Invitrogen. 2009 KPL, Inc. All rights reserved.

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Cy3 and Cy5 Conjugates Image above: Confocal fluorescence micrograph of HeLa cells stained with monoclonal antibody against mitochondria enzyme and Cy3conjugated anti-mouse antibody (red); rabbit polyclonal antibody to histones in DNA and Cy5-conjugated anti-rabbit antibody (blue).

KPL CyTM Dye Conjugates- Improve your assay and brighten your day! Cy Dyes offer intense fluorescence when coupled

photostability. Cy Dye conjugates are used in both

with KPL’s affinity purified antibodies and strepta-

visual and image analysis fluorescent microscopy

vidin. The sensitivity and reproducibility of KPL

and in situ hybridization. Cy3 conjugates are ideally

antibodies combined with the brightness of Cy3 and

used in visual color applications, whereas Cy5 con-

Cy5 dyes produce an exceptional set of conjugates

jugates emit in the far red spectrum and are not easi-

ideal for multiple labeling experiments. Dye/protein

ly visualized.

ratios have been established to ensure optimal fluorescence with minimal background. They present maximum excitation/emission spectra at 550/570 nm (Cy3) and 650/670 nm (Cy5).

Cy3

Cy5

Benefits of KPL Cy Dye Conjugates High performance conjugates – optimized dyeprotein ratios ensure high signal-to-noise. Intense fluorescence – offers greater sensitivity than TRITC conjugates.

Ex Em

Narrow emission spectra – enables sensitive, multi-color analysis.

Fluorescence

Excellent photostability – more photostable than TRITC conjugates. Consistent performance – minimal lot-to-lot variation reduces need for assay optimization. Buffer stability – after rehydration, conjugates are stable at pH 4-9. Wavelength (nm)

Cy dyes are excellent alternatives to most other fluorrescein dyes as they are brighter and offer greater

Instrument compatibility – excitation and emission spectra correspond with standard filter sets and laser settings. SEE MORE with KPL!

Excellent Performance As demonstrated below KPL Cy3- and Cy5-labeled conjugates produce brighter fluorescence than those of other suppliers.

Cy 3 and Cy5 Conjugates

Cytomegalovirus-infected cells detected with a biotinylated CMV probe and the DNADetectorTM Fluorescent in situ Hybridization Kit using Cy3-Strept-avidin and DAPI.

Microplates were coated with serially diluted mouse IgG at the indicated concentrations. Conjugates at a concentration of 0.01 mg/mL were applied and incubated for 30 minutes. Fluorescence was measured with a Perkin Elmer VICTOR 3 Multilabel Plate Reader.

Ordering Information Description Anti-Mouse IgG (γ), HSA F(ab’)2 Anti-Mouse IgG (γ), HSA Anti-Mouse IgG (H+L), HSA F(ab’)2 Anti-Mouse IgG (H+L), HSA Anti-Mouse IgG (H+L), RbSA, HSA Anti-Mouse IgM (µ), HSA Anti-Mouse IgG+IgM (H+L), HSA Anti-Rabbit IgG (H+L) F(ab’)2 Anti-Rabbit IgG (H+L), HSA Anti-Rabbit IgG (H+L), HSA Anti-Rat IgG (H+L) F(ab’)2 Anti-Human IgG (H+L) Anti-Human IgG (γ) F(ab’)2 Anti-Human IgG (γ) Anti-Human IgM (µ) F(ab’)2 Anti-Human IgM (µ) Anti-Guinea Pig IgG (H+L) Anti-Chicken IgG (H+L) Anti-Horse IgG (H+L) Anti-Swine IgG (H+L) Anti-Dog IgG (H+L) Anti-Sheep IgG (H+L) Anti-Goat IgG (H+L) Streptavidin

Cy3 072-01-18-02 202-01-18-02 072-01-18-06 202-01-18-06 072-01-18-18 072-01-18-03 072-01-18-09 072-01-15-06 202-01-15-16 072-01-15-16 072-01-16-06 202-01-10-06 072-01-10-02 202-01-10-02 072-01-10-03 202-01-10-03 072-01-17-06 072-01-24-06 072-01-21-06 072-01-14-06 072-01-19-06 072-01-23-06 072-01-13-06 072-01-30-00

Cy5 072-02-18-02 202-02-18-02 072-02-18-06 202-02-18-06 072-02-18-18 072-02-18-03 072-02-18-09 072-02-15-06 202-02-15-16 072-02-15-16 072-02-16-06 202-02-10-06 072-02-10-02 202-02-10-02 072-02-10-03 202-02-10-03 072-02-17-06 072-02-24-06 072-02-21-06 072-02-14-06 072-02-19-06 072-02-23-06 072-02-13-06 072-02-30-00

Signal Detection Cy3 is excited maximally at 550 nm and fluoresces maximally at 570 nm. It is excited to about 50% of maximum with an argon laser (514 nm or 528 nm lines), or to about 75% of maximum with a helium/neon laser (543 nm line) or mercury lamp (546 nm line). Cy5 is excited maximally at 650 nm and fluoresces maximally at 670 nm. It is excited to about 98% of maximum with a krypton/argon laser (647 nm line) or to about 63% of maximum with a helium/neon laser (633 nm line). Cy5 produces minimal autofluorescence of biological specimens in this region of the spectrum. A confocal microscope equipped with the appropriate laser for excitation and a far-red detector enable double labeling with Cy3 and Cy5.

Gaithersburg, MD 20878

HSA=human serum adsorbed RbSA=rabbit serum adsorbed Cy Dye antibody conjugates are made in goat except anti-goat and anti-sheep antibodies made in rabbit. Supplied in 1 mg lyophilized form.

Phone: 800.638.3167 Fax: 301.948.0169 www.kpl.com ISO 9001:2000 Registered

To order or for more information on KPL’s line of unlabeled and conjugated affinity purified antibodies, contact us at 800.638.3167 / 301.948.7755, fax 301.948.0169 or visit us at www.kpl.com. ML368-01 For research use only. Cy Dye is a trademark of GE Healthcare. ©2009 KPL, Inc. All rights reserved.

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Peroxidase Conjugates KPL Peroxidase Conjugates: Time-tested, Sensitive and Reliable Since 1979 KPL has provided quality affinity

streptavidin. They are affinity purified and in

purified antibodies to researchers worldwide.

some cases further adsorbed to minimize cross-

Over the years we have refined our production

reactivity between animal species or shared

process to provide antibodies with high potency

reactivity with other immunoglobulin classes.

and consistent performance in immunoassays.

HRP-labeled F(ab’)2 fragment antibodies are

From the start KPL gives careful consideration

offered for assays requiring extremely low back-

to immunogen preparation, using a highly

ground and absence of F(c)-mediated binding.

purifed formulation to generate antiserum. KPL pools antiserum from multiple animals to

KPL reacts HRP of the highest quality with

reduce natural animal to animal serum variabili-

affinity purified antibodies and streptavidin

ty. During the purification process our ISO

using the periodate method of Nakane and

9001:2008-certified quality procedures impose

Kawaoi. Special features of HRP include:

rigorous standards for potency and cross-reac-

• faster catalytic rate than alkaline phosphatase • generates more product in shorter incubation

tivity. The result is standardized antibody lots with excellent reproducibility. Our extensive line of peroxidase (HRP) conjugates is available across a range of animal species, including human, mouse, rabbit and rat antibodies, as well as other animal species and

times

• provides maximum sensitivity, low nonspecific binding

• ideal for ELISA, Western blotting and immunohistology applications. SEE MORE with KPL!

Peroxidase Conjugates Peroxidase Conjugates Ordering Information (Partial listing) Catalog#

Description

Size

KPL offers a range of sensitive substrates for use with HRP conjugates. They provide a choice of intense colors for ELISA, blotting and cell staining applications.

04-10-06

HRP-labeled Goat Anti-Human IgG (H+L)

0.1 mg

04-10-17

HRP-labeled Goat Anti-Human IgA+IgG+IgM (H+L), MSA

0.1 mg

04-10-20

HRP-labeled Goat Anti-Human IgG (Fc)

0.1 mg

074-1002

HRP-labeled Goat Anti-Human IgG (γ)

1.0 mg

074-1003

HRP-labeled Goat Anti-Human IgM (μ)

1.0 mg

074-1004

HRP-labeled Goat Anti-Human IgE (ε)

1.0 mg

074-1006

HRP-labeled Goat Anti-Human IgG (H+L)

1.0 mg

074-1007

HRP-labeled Goat Anti-Human IgA+IgG+IgM (H+L)

1.0 mg

14-10-01

HRP-labeled Goat Anti-Human IgA (α)

0.5 mg

214-1002

HRP-labeled F(ab’)2 Goat Anti-Human IgG (γ)

0.5 mg

214-1003

HRP-labeled F(ab’)2 Goat Anti-Human IgM (μ)

0.5 mg

214-1006

HRP-labeled F(ab’)2 Goat Anti-Human IgG (H+L)

0.5 mg

474-1002

HRP-labeled Goat Anti-Human IgG (γ), Liquid

1.0 mL

474-1003

HRP-labeled Goat Anti-Human IgM (μ), Liquid

1.0 mL

474-1006

HRP-labeled Goat Anti-Human IgG (H+L), Liquid

1.0 mL

•

4 CN Peroxidase Substrate

04-18-06

HRP-labeled Goat Anti-Mouse IgG (H+L), HSA

0.1 mg

•

04-18-15

HRP-labeled Goat Anti-Mouse IgG (H+L), RtSA, HSA

0.1 mg

TMB Membrane Peroxidase Substrate

04-18-18

HRP-labeled Goat Anti-Mouse IgG (H+L), RbSA, HSA

0.1 mg

074-1802

HRP-labeled Goat Anti-Mouse IgG (γ), HSA

1.0 mg

074-1803

HRP-labeled Goat Anti-Mouse IgM (μ), HSA

1.0 mg

074-1806

HRP-labeled Goat Anti-Mouse IgG (H+L), HSA

1.0 mg

074-18-061

HRP-labeled Goat Anti-Mouse IgG (H+L), XSA

1.0 mg

074-1807

HRP-labeled Goat Anti-Mouse IgA+IgG+IgM (H+L), HSA

1.0 mg

074-1809

HRP-labeled Goat Anti-Mouse IgG+IgM (H+L), HSA

1.0 mg

14-18-01

HRP-labeled Goat Anti-Mouse IgA (α), HSA

0.5 mg

214-1802

HRP-labeled F(ab’)2 Goat Anti-Mouse IgG (γ), HSA

0.5 mg

214-1806

HRP-labeled F(ab’)2 Goat Anti-Mouse IgG (H+L), HSA

0.5 mg

474-1802

HRP-labeled Goat Anti-Mouse IgG (γ), HSA, Liquid

1.0 mL

474-1806

HRP-labeled Goat Anti-Mouse IgG (H+L), HSA, Liquid

1.0 mL

074-1506

HRP-labeled Goat Anti-Rabbit IgG (H+L)

1.0 mg

074-15-061

HRP-labeled Goat Anti-Rabbit IgG (H+L), XSA

1.0 mg

074-1516

HRP-labeled Goat Anti-Rabbit IgG (H+L), HSA

1.0 mg

214-1516

HRP-labeled F(ab’)2 Goat Anti-Rabbit IgG (H+L), HSA

0.5 mg

474-1506

HRP-labeled Goat Anti-Rabbit IgG (H+L), Liquid

1.0 mL

474-1516

HRP-labeled Goat Anti-Rabbit IgG (H+L), HSA, Liquid

1.0 mL

14-30-00

HRP-labeled Streptavidin

0.5 mg

Phone: 800.638.3167

474-3000

HRP-labeled Streptavidin, Liquid, Molecular Grade

1.0 mL

Fax: 301.948.0169

Visit our website at www.kpl.com for a complete listing of HRP-labeled antibodies. To order or for more information on KPL’s protein research products, contact us at 800.638.3167 / 301.948.7755, FAX 301.948.0169 or visit us at www.kpl.com.

ELISA •

ABTS® 1- and 2-Component Microwell Peroxidase Substrates

•

SureBlueTM TMB Peroxidase Substrate

•

SureBlue ReserveTM TMB Peroxidase Substrate

•

TMB Peroxidase Substrate

Blotting

•

LumiGLO®Chemiluminescent Substrate LumiGLO ReserveTM Chemiluminescent Substrate

Whichever substrate you choose, enjoy the benefits of excellent signalto-noise and reproducibility.

Gaithersburg, MD 20878

www.kpl.com

ISO 9001:2008 Registered

SEE MORE with KPL!

Ordering Information

Catalog #

Description

Size

Protein Detector™ Western Blotting Kits Each kit includes anti-mouse and anti-rabbit conjugates, Detector Block, Wash Solution Concentrate and Substrate. Phosphatase Chromogenic 55-11-50

BCIP/NBT Western Blot Kit

2500 cm2

Peroxidase Chromogenic 54-11-50

TMB Western Blot Kit

Peroxidase Chemiluminescent 54-12-50 LumiGLO® Western Blot Kit 54-13-50

LumiGLO Reserve Blot Kit

Western

2500

cm2

2500 cm2 2400

cm2

Related Reagents and Kits Antibody Conjugates All antibodies listed below are produced in goat. For a complete antibody listing, refer to KPL’s Product Catalog.

Catalog #

Description

Size

Substrates for Western Blotting Phosphatase Colorimetric Substrates 50-81-18 BCIP/NBT Substrate

100 mL

50-81-07

BCIP/NBT Substrate

600 mL

50-81-30

FirePhosTM Membrane AP Substrate

100 mL

50-81-40

FirePhos Membrane AP Substrate

400 mL

50-81-34

FirePhos Membrane AP Substrate

1000 mL

Phosphatase Chemiluminescent Substrates 55-60-03 PhosphaGLOTM AP Substrate

30 mL

55-60-04

PhosphaGLO AP Substrate

100 mL

55-60-01

PhosphaGLO Reserve AP Substrate

30 mL

55-60-02

PhosphaGLO Reserve AP Substrate

100 mL

Peroxidase Chromogenic Substrates 50-77-18 TMB Membrane Substrate

100 mL

50-77-03

TMB Membrane Substrate

200 mL

Phosphatase-labeled 475-1006 Anti-Human IgG (H+L)

1.0 mL, liquid

50-73-00

4 CN Substrate

600 mL

475-1806

Anti-Mouse IgG (H+L), HSA

1.0 mL, liquid

50-73-04

4 CN Substrate

2700 mL

475-1506

Anti-Rabbit IgG (H+L)

1.0 mL, liquid

Peroxidase Chemiluminescent Substrates 54-61-02 LumiGLO Chemiluminescent Substrate

60 mL

Peroxidase-labeled 474-1006 Anti-Human IgG (H+L)

1.0 mL, liquid

54-61-00

LumiGLO Chemiluminescent Substrate

240 mL

474-1806

Anti-Mouse IgG (H+L) HSA

1.0 mL, liquid

54-61-01

LumiGLO Chemiluminescent Substrate

720 mL

474-1506

Anti-Rabbit IgG (H+L)

1.0 mL, liquid

54-71-00

LumiGLO Reserve Substrate Kit

2400 cm2

54-71-01

LumiGLO Reserve Substrate Kit

600 cm2

Biotin-labeled 16-10-06 Anti-Human IgG (H+L)

0.5 mg

176-1006

2.0 mg

Anti-Human IgG (H+L)

Labeled Streptavidin 474-3000 HRP-labeled 475-3000

AP-labeled

Assay Support Reagents 50-84-00 Coating Solution Concentrate

50 mL

54-15-01

HRPStabilizer

200 mL

1.0 mL, liquid

55-15-00

APStabilizer

200 mL

1.0 mL, liquid

50-61-00

10% BSA Diluent/Blocking Solution Concentrate

200 mL

50-82-01

Milk Diluent/Blocking Solution Concentrate

200 mL

71-83-00

DetectorTM Block (5X)

240 mL

50-63-00

Wash Solution Concentrate (20X)

800 mL

50-63-06

Biotin Wash Solution Concentrate (10X) 200 mL

60-00-50

Biodyne® B Nylon Membrane

1 roll

HSA=human serum adsorbed.

To order or for more information, call KPL at 800.638.3167, 301.948.7755, Fax: 301.948.0169. www.kpl.com or contact your local sales partner. ML282-04

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