Near-IR CF® Dye conjugates for western blotting
Near-infrared (near-IR) western detection is highly sensitive, and offers advantages of wider linear range and multiplexing capability compared to chemiluminescence detection (see our webinar to learn more). Biotium’s near-IR CF® dyes are the brightest and most photostable available. Learn more about CF®680 and CF®770 dyes for near-IR western, In-Cell Western®, and other applications.
We offer wide selection of primary and secondary antibodies conjugated to our exceptional near-IR CF® dyes for western blot. We also offer HRP conjugates for chemiluminescence detection.
![](https://biotium.com/wp-content/uploads/2017/01/CF680-CF770-vs-IR680LT-IR800CW-389x275.jpg)
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Near-IR CF Dye Secondary Antibodies for Multiplex WB
Conjugate | Cross-adsorption | CF®680 (681/698 nm) | CF®680R (680/701 nm) | CF®750 (755/777 nm) | CF®770 (770/797 nm) | CF®790 (784/806 nm) |
---|---|---|---|---|---|---|
Donkey Anti-Goat | Ck, GP, Hs, Hu, Ms, Rb, Rt, SHm | 20060 | 20196 | 20362 | 20277 | |
Donkey Anti-Guinea Pig | Bv, Ck, Gt, Hs, Hu, Ms, Rb, Rt, Shp, SHm | 20241 | 20242 | |||
Donkey Anti-Mouse | Bv, Ck, Gt, GP, Hs, Hu, Rb, Shp, SHm | 20194 | 20363 | |||
Donkey Anti-Rabbit | Bv, Ck, Gt, GP, Hs, Hu, Ms, Rt, Shp, SHm | 20418 | 20195 | 20298 | 20484 | 20344 |
Donkey Anti-Sheep | Ck, GP, Hs, Hu, Ms, Rb, Rt, SHm | 20062 | ||||
Goat Anti-Guinea Pig | Bv, Ck, Gt, Hs, Hu, Ms, Rb, Rt, SHm, Shp | 20497 | 20496 | 20499 | 20498 | |
Goat Anti-Mouse | Bv, Hs, Hu, Rb, Sw | 20065 | 20192 | 20463 | 20077 | 20342 |
Goat Anti-Mouse IgG1 | Bv, Hu, Rb | 20253 | 20254 | |||
Goat Anti-Mouse IgG2a | Bv, Hu, Rb | 20263 | 20842 | 20264 | ||
Goat Anti-Mouse IgG2b | Bv, Hu, Rb | 20273 | 20430 | 20274 | ||
Goat Anti-Rabbit | Hu, Ms, Rt | 20067 | 20193 | 20078 | 20343 | |
Goat Anti-Rat | Bv, Hs, Hu, Rb | 20069 | 20383 | |||
Rabbit Anti-Mouse | Hu | 20061 |
Bv: bovine, Ck: chicken, GP: guinea pig, Gt: goat, Hs: horse, Hu: human, Ms: mouse, Rb: rabbit, Rt: rat, SHm: Syrian hamster, Shp: sheep, Sw: swine
TrueBlack® WB Blocking Buffer Kit
The TrueBlack® WB Blocking Buffer Kit is a ready-to-use buffer system for fluorescence-based western blotting (WB). The buffers yield optimal specificity and sensitivity by blocking non-specific interactions of dye-labeled antibodies with proteins and the blotting membrane.
TrueBlack® WB Blocking Buffer Kit Features
- Blocks as well or better than Odyssey® Blocking Buffer, at a lower price
- Reduces non-specific protein bands and background over entire membrane
- Suppresses background from charged dyes better than BSA, gelatin, or casein
- Compatible with PVDF and nitrocellulose membranes
- Contains no mammalian proteins, for broad antibody compatibility
- For visible and near-IR fluorescent westerns
![](https://biotium.com/wp-content/uploads/2018/07/TrueBlack-WB-phosphoprotein-composite-193x275.jpg)
Superior western blocking for next-generation fluorescent dyes
Non-specific signal in WB can arise from multiple sources, including antibody cross-reactivity with off-target proteins, non-specific antibody adsorption to the membrane, and membrane autofluorescence. Another potential cause of background is the effect of fluorescent dyes themselves on the specificity of labeled antibodies. Highly charged dyes like Alexa Fluor® or CF® dyes have improved solubility and brightness of conjugates compared to uncharged dyes. However, the extra charge carried by antibodies labeled with these dyes can result in non-specific binding to proteins and membranes. The TrueBlack® WB Blocking Buffer Kit blocks background from multiple sources including charged dye conjugates (Figure 2). TrueBlack® blocking buffer is especially advantageous for phosphoprotein detection, significantly improving specificity compared to conventional blocking buffers (Figure 1).
Switch from Odyssey® Blocking Buffer and save
TrueBlack® WB Blocking Buffer performs as well or better for fluorescent WB compared to LI-COR’s Odyssey® Blocking Buffer (Figure 1), and is priced lower on a per membrane basis.
![](https://biotium.com/wp-content/uploads/2018/07/TrueBlack-WB-tubulin-composite-293x275.jpg)
Compare TrueBlack® WB Blocking Buffer Kit with Odyssey® Blocking Buffer
Product | TrueBlack® WB Blocking Buffer Kit | Odyssey® Blocking Buffer |
---|---|---|
Trial Size | For 10 membranes | 125 mL for 4 membranes |
Full Size | For 50 membranes | 500 mL for 16 membranes |
VIEW & PURCHASE
VersaBlot™ Near-IR Total Protein Normalization Kits
VersaBlot™ Total Protein Normalization Kits allow simple, sensitive and highly linear protein quantitation on SDS-PAGE gels and western blot membranes. The kits allow you to label purified proteins or cell lysates with our near-infrared CF® dyes before running the samples on SDS-PAGE. Proteins can then be visualized on a gel or membrane using a fluorescent gel scanner, allowing detection of as little as 1 ng protein per band. If desired, the prestain can be reversed after scanning, facilitating downstream multi-color western blot analysis. The staining demonstrates excellent linearity for quantitation of total protein over a wide dynamic range, outperforming traditional western blot normalization based on housekeeping protein detection.
VersaBlot™ Total Protein Prestain Features
- Reversible prestain for downstream multi-color western blot analysis
- Superior linearity for western normalization compared to housekeeping proteins
- Highly sensitive protein quantitation on PAGE gels or western membranes
- Fast and simple labeling of proteins or lysates, no purification required
- Detect as little as 1 ng protein and 10% difference in protein content
![](https://biotium.com/wp-content/uploads/2017/12/MnS-Total-Protein-Prestain-CF680T-BSA-gel-388x275.png)
![](https://biotium.com/wp-content/uploads/2017/12/MnS-Total-Protein-Prestain-CF770T-lysate-blot-400x275.png)
Product Information
Product | Catalog number | Size | Abs/Em | Imaging Systems / Detection channels |
---|---|---|---|---|
VersaBlot™ CF®680T Total Protein Normalization Kit | 33025-T | 100 labelings | 681 / 698 nm | Amersham Typhoon™ Trio Cy®5 channel Amersham Typhoon™ 5 IR short channel LI-COR® Odyssey® 700 channel |
33025 | 500 labelings | |||
VersaBlot™ CF®770T Total Protein Normalization Kit | 33026-T | 100 labelings | 764 / 787 nm | Amersham Typhoon™ 5 IR long channel LI-COR® Odyssey® 800 channel |
33026 | 500 labelings |
Typhoon is a trademark and Cy dye is a registered trademark of Cytiva; Odyssey is a registered trademark of LI-COR Biosciences.
RedDot™1 Far-Red Nuclear Stain for In-Cell Western®
RedDot™1 is a far-red cell membrane-permeable nuclear dye similar to Draq5™. Staining does not require a wash step and demonstrates greater photostability than the traditional blue fluorescent nuclear stains DAPI and Hoechst. RedDot™1 can be excited at a wide range of wavelengths between 488 nm and 647 nm, with an emission maximum at 694 nm. RedDot™1 is particularly useful for cell number normalization for In-Cell Western® assays using a near-infrared scanner such as the LI-COR® Odyssey®. In fixed cells, RedDot™ 1 staining generates a highly linear fluorescence signal in the 700 nm channel that is proportional to the number of cells. In a direct comparison with DRAQ5™/Sapphire700™, RedDot™ 1 staining demonstrates a higher signal and a wider linear range.
RedDot™1 Features
- Ideal for normalization of In-Cell Western® in fixed cells, similar to Draq5™/Sapphire®700
- Highly thermostable and photostable, for convenient handling and demanding imaging applications
- Can also be used for DNA content analysis by flow cytometry like Vybrant™ DyeCycle™ Ruby
- λEx/λEm = 662/694 nm (with DNA), for detection in the Cy®5 channel
RedDot™1 staining of HeLa cells for cell number normalization. HeLa cells were seeded in 96 wells at the indicated densities. After 24 hours, cells were fixed, permeabilized, and stained with the indicated dyes for one hour at room temperature according to the supplier’s protocol for DRAQ5™/Sapphire700™. Linearity of fluorescence signal for 0-6250 cells per well (left). Linearity of fluorescence signal for 0-50,000 cells per well (right). HeLa cells seeded at 25,000 cells per well were confluent at the time of assay.
Product | Catalog No. | Size | Color (Ex/Em) | Applications |
---|---|---|---|---|
RedDot™1 Far-Red Nuclear Stain, 200X in Water | 40060-T | 25 uL | (Far-red) 662/694 nm | • Live cell nuclear stain comparable to Draq5™ • Ideal for In-Cell Western® normalization • Suitable for microscopy and cell cycle analysis by flow cytometry |
40060 | 250 uL | |||
40060-1 | 1 mL |
Chromogenic Detection
Chromogenic detection by horseradish peroxidase (HRP) or alkaline phosphatase is a widely used and cost-effective method for western blots analysis. Colorimetric detection works by the production of an insoluble colored byproduct from a chemical reaction between a substrate and reporter enzyme. Biotium offers several secondary and anti-tag antibodies conjugated to either HRP or alkaline phosphatase for chromogenic detection. See the tables below for available conjugates. You can also view our entire Secondary Antibody Product Listings for other HRP and alkaline phosphatase conjugates including anti-human IgG, IgA, IgM antibodies. Biotium also offers several colorimetric substrates for alkaline phosphatase, and a DAB substrate kit for HRP listed in the table below.
HRP and Alkaline Phosphatase Secondary Antibodies
Conjugate | Donkey Anti-Mouse (Min X Rat) | Donkey Anti-Rabbit | Goat Anti-Chicken | Goat Anti-Mouse | Goat Anti-Rabbit | Goat Anti-Rat | Chicken Anti-Goat | Goat Anti-Llama | Goat Anti-Mouse | Goat Anti-Rabbit |
---|---|---|---|---|---|---|---|---|---|---|
Cross-Adsorption | Bv, Ck, Gt, GP, Hs, Hu, Rb, Rt, Shp, SHm | Bv, Ck, Gt, GP, Hs, Hu, Ms, Rt, Shp, SHm | Bv, Gt, GP, Hs, Hu, Ms, Rb, Rt, SHm, Shp | Bv, Hs, Hu, Rb, Sw | Hu, Ms, Rt | Bv, Hs, Hu, Rb | None | None | None | None |
HRP | 20404 | 20405 | 20474 | 20401 | 20403 | 20406 | 20839 | 20475 | 20400 | 20402 |
Alkaline Phosphatase | 20466 | 20467 | 20464 | 20465 |
Bv: bovine, Ck: chicken, GP: guinea pig, Gt: goat, Hs: horse, Hu: human, Ms: mouse, Rb: rabbit, Rt: rat, SHm: Syrian hamster, Shp: sheep, Sw: swine
HRP and Alkaline Phosphatase Anti-Tag Antibodies
Don’t see what you’re looking for? Contact us! We may be able to add a new conjugate to our catalog, or perform a custom labeling for you.Chromogenic Substrates for HRP and Alkaline Phosphatase
Product | Catalog No. | Size | Substrate for | Precipitate Color |
---|---|---|---|---|
BCIP, NA | 1001 | 100 mg | Alkaline Phosphatase | Dark Blue |
1001-1 | 500 mg | |||
1001-2 | 5 g | |||
BCIP, Toluidine | 10002 | 100 mg | Alkaline Phosphatase | Dark Blue |
10002-1 | 500 mg | |||
10002-2 | 5 g | |||
BCIP Pink | 10006 | 100 mg | Alkaline Phosphatase | Pink |
BCIP Pink/NBT Kit | 10007 | 1 set | Alkaline Phosphatase | Pink |
BCIP Red | 10004 | 100 mg | Alkaline Phosphatase | Red |
BCIP Red/NBT Kit | 10005 | 1 set | Alkaline Phosphatase | Red |
BCIP/NBT Kit | 10003 | 1 set | Alkaline Phosphatase | Dark Blue |
Chromogenic Phosphatase Substrate Sampler | 10022 | 1 kit | Alkaline Phosphatase | Multiple |
DAB Substrate Kit | 30015 | 1 kit | HRP | Brown |
Accessory Reagents for Western Blotting
See our selection of prestained markers, buffers, blocking agents, detergents, and other reagents and accessories below.
Product Information
Product | Catalog No. | Size | Features |
---|---|---|---|
Peacock™ Prestained Protein Marker | 21530 | 50 uL or 500 uL | • Visible color protein marker for SDS-PAGE and western • 8 blue bands ranging from 10 kDa to 180 kDa • Red/green bands at 75 kDa/25 kDa |
Peacock™ Plus Prestained Protein Marker | 21531 | 50 uL or 500 uL | • Visible color protein marker for SDS-PAGE and western • 10 blue bands ranging from 8 kDa to 245 kDa • Red/green bands at 75 kDa/25 kDa |
4X Protein Loading Buffer with Orange Tracking Dye | 40136 | 15 mL | • A convenient 4X sample loading buffer for protein gel electrophoresis • Optimized for use with denaturing SDS-PAGE • Orange tracking dye avoids unwanted background fluorescence caused by blue tracking dyes |
Ponceau S Solution | 22001 | 1 L | • Stain proteins on PVDF or nitrocellulose membranes with visible pink dye • Fast & reversible visualization of protein transfer before western detection |
TrueBlack® WB Blocking Buffer Kit | 23013 | For 10 membranes or 50 membranes | • Superior blocking for fluorescent WB • Works as well or better than Odyssey® Blocking Buffer, at a lower cost • Suppresses background caused by charged fluorescent dyes • Reduces antibody cross-reactivity, eliminating non-specific bands |
10X Fish Gelatin Blocking Agent | 22010 | 100 mL | • Provides excellent blocking for IF or western • Add to buffer of your choice (PBS or TBS) • Compatible with anti-goat and anti-sheep secondaries |
Fish Gelatin Powder | 22011 | 2 x 50 g | • Gelatin from cold water fish skin for blocking for IF or western • Compatible with anti-goat and anti-sheep secondaries |
Bovine Serum Albumin, 30% Solution | 22014 | 100 mL | • Commonly used blocking agent and antibody or protein stabilizer • 30% solution in water • Made from IgG-free, protease-free Fraction V BSA |
Bovine Serum Albumin Fraction V | 22013 | 50 g | • Commonly used blocking agent and antibody or protein stabilizer • IgG-free, protease-free Fraction V BSA |
Dry Milk Powder | 22012 | 4 x 25 g | • Nonfat dry milk • Commonly used blocking agent for western |
Tween®-20 | 22002 | 50 mL | • Detergent commonly used for western blocking and washing |
Mini Cell Scrapers | 22003 | Pack of 200 | • For harvesting cells or cell lysates from 96-, 48- and 24-well plates • 0.5 cm (3/16″) wide and 6 cm (2 3/8″) long • 20 packs of 10 scrapers per pack • Polyethylene, disposable & sterile |
Ultrafiltration Vials (3K MWCO) | 22018 | Pack of 5 | • For removing buffers, salts, and free dyes from proteins or DNA • Simple microcentrifuge spin-column format • 3 kDa molecular weight cut-off (MWCO) |
Ultrafiltration Vials (10K MWCO) | 22004 | Pack of 5 | • For removing buffers, salts, and free dyes from proteins or DNA • Simple microcentrifuge spin-column format • 10 kDa molecular weight cut-off (MWCO) |
DTT | 91050 | 1 g | • Reducing agent commonly used to prepare samples for SDS-PAGE |
TCEP | 91049 | 1 g | • Odorless reducing agent • More effective and stable than DTT |
![](https://biotium.com/wp-content/uploads/2018/01/Peacock-and-Peacock-Plus-159x275.jpg)