-Histofine®

DAB-2VNICHIREI BIOSCIENCES INC.
6-19-20, Tsukiji, Chuo-ku, Tokyo, 104-8402, JAPAN
Phone: 81-3-3248-2207 Facsimile: 81-3-3248-2243

Peroxidase Chromogen/Substrate solution

3,3’-DIAMINOBENZIDINE TETRAHYDROCHLORIDE (DAB) SUBSTRATE SOLUTION
SUBSTRATE FOR HORSERADISH PEROXIDASE
Package/Code

500 tests/425312F, 1500 tests/425314F

FORM
Liquid.
This product consists of followings:
Reagent A: DAB solution concentrate
Reagent B: DAB buffer
Tube
Chip (For Reagent B)
Dropping bottle…
…
…
…
…

[500 tests]
1.2mL×230.0mL×2×1×1×1[1500 tests]
1.2mL×630.0mL×6×1×1×1

PREPARATION of Chromogen/Substrate solution
Add 1 drop (approx. 20μL) of Reagent A Chromogen reagent per 500μL of Reagent B Chromogen substrate into graduatedtube, and place its cap then mix carefully without foaming.
Remove full dosage of the substrate solution from the tube to dropping bottle using pipette as well for use.
Store in light shielded condition at 2 to 8°C and use within 2 weeks after the preparation.
Note: 1. Use the substrate solution removing foams or exclusive of foams when its surface is foamed by mixing.
2. Do not use mixture of substrate solutions prepared different day.

USAGE
Brownish-red precipitates are produced by the reaction of this substrate solution with peroxidase. Such precipitates are
insoluble in organic solvent.
Adjust the substrate solution to room temperature prior to use.
React the objective tissue section with peroxidase conjugate in advance.
At the same time, staining of negative control slide using such as PBS in place of primary antibody is recommended to beprepared for checking the staining result.
Chip
Reagent B is provided with translucent chip placed on its bottle. However the chip is not allowed for applying solution
drop by drop, therefore it is not suitable for small volume preparation. For such preparation, replace the translucent chipby colored chip provided exclusively for dropping is recommended. The colored chip is suitable for applying solution
drop by drop.
Dropping bottle
Dropping bottle is used for storage of reagents, not only for dropping of substrate solution.
Reuse
Tube and dropping bottle are reusable after wash with purified water and dry completely.

STAINING PROCEDURE
Apply the substrate solution using dropping bottle covering the tissue section on the slide completely and incubate at room
temperature for 5 to 20 minutes after addition, reaction and washing of peroxidase conjugate.
For chromogenic condition, adjust reaction time by microscopic observation comparing with negative control slide.
After adequate chromogenic reaction, wash the slide well with tap water.
Mount the slide with permanent mounting media and fix with cover slip after dehydrated in graded series of alcohol andcleared in xylene.

STORAGEStore at 2-8°C.

REMARKSFor research use only. Not for diagnostic use.
11/2015WARNING and PRECAUTIONS
1. Avoid contact with eyes, skin and clothing as well as ingestion and inhalation.
2. Handle Reagent A with adequate attention due to its DAB evolves into mutagens is recognized.
3. Handle Reagent B with adequate attention due to hydrogen peroxide solution contained.
4. Dispose unused Reagent A and substrate solution properly in accordance with all applicable rules and regulations oflocal and country due to its DAB contained.

-Histofine is NICHIREI BIOSCIENCES’s trademark. The registration countries would be referred to us.

11/2015