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281751 DAB, Tetrahydrochloride, 50X Concentrate

281751
Purchase on Sigma-Aldrich

Overview

Replacement Information

Key Spec Table

Products

Catalogue NumberPackaging Qty/Pack
281751-10ML Glass bottle 10 ml
Description
OverviewProduces a brown alcohol-insoluble end product. Also useful as a stain for myelin in glutaraldehyde-fixed sections. Supplied as a 50X concentrate.
Catalogue Number281751
Brand Family Calbiochem®
Synonyms3,3ʹ-Diaminobenzidine
References
ReferencesKrueger, S.K., et al. 1999. Biotech. Histochem. 74, 105.
Larrson, L.I. 1988. Immunocytochemistry: Theory and Practice, CRC Press, Inc., Boca Raton, FL.
Gallyas, F., et al. 1982. J. Histochem. Cytochem. 30, 183.
Hsu, S.M. and Soban, E. 1982. J. Histochem. Cytochem. 30, 1079.
Lewis, P.R. and Knight, D.P. 1977. Staining Methods for Sectioned Material, In Practical Methods in Electron Microscopy, A.M. Glauert, ed. North-Holland, Amsterdam.
Graham, R.C., Jr. and Karnovsky, M.J. 1966. J. Histochem. Cytochem. 14, 291.
Product Information
CAS number7411-49-6
FormBrown to dark brown solution
FormulationSupplied as a 50X concentrate.
PreservativeNone
Quality LevelMQ100
Applications
Application NotesImmunoblotting (see comments)
Immunohistochemistry (see comments)
Application CommentsStable at 4°C and 18-26°C.

Suggested Procedure for Immunohistochemical Staining Using DAB:

1. Complete all required incubations with antibodies and HRP-labeled probes, and all necessary wash steps.
2. Prepare 1X DAB Substrate Buffer: Add 9 parts distilled water to 1 part 10X DAB Substrate Buffer (Cat. No. 281753). Mix well. [Note: 1 M Tris-HCl, pH 7.6 may be substituted for Cat. No. 281753, such that the final (1X) concentration is 100 mM Tris-HCl, pH 7.6.
3. Prepare 1X DAB Substrate Solution: Add 1 part 50X DAB Concentrate and 1 part 0.5% H2O2 to 50 parts 1X DAB Substrate Buffer (prepared in Step 2). Mix well and protect from light.
4. Following the final wash, completely cover tissue sections with the 1X DAB Substrate Solution (prepared in Step 3) and incubate 5-15 min at room temperature. Note: Variables associated with assay conditions will dictate the proper reaction time.
5. After reaction is complete, wash tissue sections thoroughly in distilled water.
6. Counterstain with hematoxylin if desired.
7. Dehydrate in graded alcohols, xylene, or xylene substitutes.
8. Mount tissue sections using xylene-based mounting media.

Suggested Procedure for Immunoblot Staining with DAB:

1. Complete all required incubations with antibodies and HRP-labeled probes, and all necessary wash steps.
2. Prepare 1X DAB Substrate Buffer: Add 9 parts distilled water to 1 part 10X DAB Substrate Buffer (Cat. No. 281753). Mix well. [Note: 1 M Tris-HCl, pH 7.6 may be substituted for Cat. No. 281753, such that the final (1X) concentration is 100 mM Tris-HCl, pH 7.6.]
3. Prepare 1X DAB Substrate Solution: Add 1 part 50X DAB Concentrate and 1 part 0.5% H2O2 to 50 parts 1X DAB Substrate Buffer (prepared in Step 2). Mix well and protect from light.
4. Following the final wash, completely cover membranes with 1X DAB Substrate Solution (prepared in Step 3). Incubate 5-30 min at room temperature.
Note: Variables associated with assay conditions will dictate the proper reaction time.
5. After reaction is complete, wash membranes thoroughly in distilled water.
6. Air-dry membranes and store protected from light.
Biological Information
Physicochemical Information
Dimensions
Materials Information
Toxicological Information
Safety Information according to GHS
RTECSDV8753000
Safety Information
Product Usage Statements
Storage and Shipping Information
Ship Code Ambient Temperature Only
Toxicity Toxic
Hazardous Materials Attention: Due to the nature of the Hazardous Materials in this shipment, additional shipping charges may be applied to your order. Certain sizes may be exempt from the additional hazardous materials shipping charges. Please contact your local sales office for more information regarding these charges.
Storage +2°C to +8°C
Do not freeze Ok to freeze
Special InstructionsDiscard if a precipitate forms or if reagent is purple (the purple material is a DAB oxidation product that binds avidly to heme-containing proteins).
Packaging Information
Transport Information
Supplemental Information
Specifications

Documentation

DAB, Tetrahydrochloride, 50X Concentrate SDS

Title

Safety Data Sheet (SDS) 

DAB, Tetrahydrochloride, 50X Concentrate Certificates of Analysis

TitleLot Number
281751

References

Reference overview
Krueger, S.K., et al. 1999. Biotech. Histochem. 74, 105.
Larrson, L.I. 1988. Immunocytochemistry: Theory and Practice, CRC Press, Inc., Boca Raton, FL.
Gallyas, F., et al. 1982. J. Histochem. Cytochem. 30, 183.
Hsu, S.M. and Soban, E. 1982. J. Histochem. Cytochem. 30, 1079.
Lewis, P.R. and Knight, D.P. 1977. Staining Methods for Sectioned Material, In Practical Methods in Electron Microscopy, A.M. Glauert, ed. North-Holland, Amsterdam.
Graham, R.C., Jr. and Karnovsky, M.J. 1966. J. Histochem. Cytochem. 14, 291.
Data Sheet

Note that this data sheet is not lot-specific and is representative of the current specifications for this product. Please consult the vial label and the certificate of analysis for information on specific lots. Also note that shipping conditions may differ from storage conditions.

Revision21-August-2007 JSW
Synonyms3,3ʹ-Diaminobenzidine
ApplicationImmunoblotting (see comments)
Immunohistochemistry (see comments)
DescriptionPeroxidase substrate that forms an insoluble, brown precipiate. Supplied as a 50X concentrate. Designed for use with DAB Substrate Buffer (Cat. No. 281753).
BackgroundSince first introduced by Graham and Karnovsky numerous procedures for the use of DAB for detection of horseradish peroxidase (HRP)-labeled probes in histochemistry, immunohistochemistry, western blots and dot blots have been described. In the presence of horseradish peroxidase and hydrogen peroxide, DAB is oxidized to a brown polymer that is insoluble in most organic solvents. Thus, xylene-based mounting media may be used for immunohistochemical applications. Sites of HRP activity on tissue sections and blots appear as brown-orange deposits. The color can be modified and intensified by treatment with salts of silver, copper, nickel, cobalt and osmium. This DAB substrate preparation is a stable, convenient, liquid concentrate in a proprietary solvent. The stabilization system prevents formation of partially oxidized DAB, thus eliminating the nonspecific binding to other heme-containing proteins so often observed with powdered DAB preparations. The concentrate can be diluted in appropriate peroxide-containing buffers, providing the researcher with the capability of formulating any of the numerous published DAB reaction systems.
FormBrown to dark brown solution
FormulationSupplied as a 50X concentrate.
CAS number7411-49-6
RTECSDV8753000
PreservativeNone
CommentsStable at 4°C and 18-26°C.

Suggested Procedure for Immunohistochemical Staining Using DAB:

1. Complete all required incubations with antibodies and HRP-labeled probes, and all necessary wash steps.
2. Prepare 1X DAB Substrate Buffer: Add 9 parts distilled water to 1 part 10X DAB Substrate Buffer (Cat. No. 281753). Mix well. [Note: 1 M Tris-HCl, pH 7.6 may be substituted for Cat. No. 281753, such that the final (1X) concentration is 100 mM Tris-HCl, pH 7.6.
3. Prepare 1X DAB Substrate Solution: Add 1 part 50X DAB Concentrate and 1 part 0.5% H2O2 to 50 parts 1X DAB Substrate Buffer (prepared in Step 2). Mix well and protect from light.
4. Following the final wash, completely cover tissue sections with the 1X DAB Substrate Solution (prepared in Step 3) and incubate 5-15 min at room temperature. Note: Variables associated with assay conditions will dictate the proper reaction time.
5. After reaction is complete, wash tissue sections thoroughly in distilled water.
6. Counterstain with hematoxylin if desired.
7. Dehydrate in graded alcohols, xylene, or xylene substitutes.
8. Mount tissue sections using xylene-based mounting media.

Suggested Procedure for Immunoblot Staining with DAB:

1. Complete all required incubations with antibodies and HRP-labeled probes, and all necessary wash steps.
2. Prepare 1X DAB Substrate Buffer: Add 9 parts distilled water to 1 part 10X DAB Substrate Buffer (Cat. No. 281753). Mix well. [Note: 1 M Tris-HCl, pH 7.6 may be substituted for Cat. No. 281753, such that the final (1X) concentration is 100 mM Tris-HCl, pH 7.6.]
3. Prepare 1X DAB Substrate Solution: Add 1 part 50X DAB Concentrate and 1 part 0.5% H2O2 to 50 parts 1X DAB Substrate Buffer (prepared in Step 2). Mix well and protect from light.
4. Following the final wash, completely cover membranes with 1X DAB Substrate Solution (prepared in Step 3). Incubate 5-30 min at room temperature.
Note: Variables associated with assay conditions will dictate the proper reaction time.
5. After reaction is complete, wash membranes thoroughly in distilled water.
6. Air-dry membranes and store protected from light.
Storage +2°C to +8°C
Do Not Freeze Ok to freeze
Special InstructionsDiscard if a precipitate forms or if reagent is purple (the purple material is a DAB oxidation product that binds avidly to heme-containing proteins).
Toxicity Toxic
ReferencesKrueger, S.K., et al. 1999. Biotech. Histochem. 74, 105.
Larrson, L.I. 1988. Immunocytochemistry: Theory and Practice, CRC Press, Inc., Boca Raton, FL.
Gallyas, F., et al. 1982. J. Histochem. Cytochem. 30, 183.
Hsu, S.M. and Soban, E. 1982. J. Histochem. Cytochem. 30, 1079.
Lewis, P.R. and Knight, D.P. 1977. Staining Methods for Sectioned Material, In Practical Methods in Electron Microscopy, A.M. Glauert, ed. North-Holland, Amsterdam.
Graham, R.C., Jr. and Karnovsky, M.J. 1966. J. Histochem. Cytochem. 14, 291.