For life science research only. 
Not for use in diagnostic procedures.
BM Purple
AP Substrate precipitating, (NBT/BCIP ready-to-use)
Cat. No. 11 442 074 001
y Version 19.0
Content version: January 2011
100 ml
Store at +2 to +8°C
F Store protected from light !
Product overview
Contents
Ready-to-use solution. Appearance prior to use: light
yellow color, after color development dark purple
Product
description
BM Purple is a chromogenic substrate for alkaline
phosphatase (AP) for precipitating enzyme immunoassays. It develops a permanent, dark purple band or
spot at the AP binding site on the membrane or solid
support. The stabilized substrate solution is be used
directly from the bottle with no mixing or reconstitution. BM Purple produces negligible background staining for more sensitivity.
Application
Storage/
stability
Additional
reagents 
required
• For washing purposes: 
PBST: Phosphate buffered saline (PBS), pH 7.2 – 7.5
containing 0.2 % Tween 20* (v/v)
• For the blocking of the membrane: 
Blocking Reagent* or PBS containing 1 – 5% bovine
or horse serum (v/v)
• Antibody-AP conjugate
Procedure
Carry out your membrane ELISA or immunoblot on
nitrocellulose according to standard protocols.
Step
Action
• Immunohisto/cytochemistry
• Western blot
• Southern blot
• Northern blot
• Colony and plaque hybridization
• In situ hybridization
These assays may be either qualitative or quantitative.
The unopened bottle is stable at +2 to +8°C until the
expiration date printed on the label. 
Note: Store protected from light!
1
Apply antigen to membrane, 1 – 2 ␮g per spot.
2
Wash membrane twice in PBST.
3
Block remaining protein binding sites with
Blocking reagent. or horse serum.
4
Wash membrane twice in PBST.
5
Incubate with antibody-AP conjugate.
6
Wash membrane twice with PBST.
7
Rinse with double dist. water.
8
Bring BM Purple Substrate to +15 to +25°C;
invert once to mix.
9
Incubate membrane in BM Purple Substrate at
+15 to +25°C.
Note: Color development may be complete in
30 min.
10
Rinse with double dist. water.
Protocol for Immunoblots
11
Handling
Instructions
Dry between sheets of filter paper or paper
towel.
12
Store in the dark.
Note: The colored reaction product is stable
when dried.
0111.11470 0430019
• Bring the substrate to +15 to +25°C before staining
and use it right out of the bottle; do not dilute it.
• Wash blots in petri dishes or shallow trays on
rotating plate shaker.
• To minimize use of reagents, prepare spot blots in
1 × 8 cm nitrocellulose strips and lay them in 10 cm
petri dishes. Carefully pipette 2 ml antibody conjugate directly onto each strip and incubate.
• Background staining should be negligible or, if
present, should fade with drying. If it persists, try
increasing the serum concentration in the blocking
buffer, increasing blocking time, diluting the conjugate, or testing a different batch of nitrocellulose.
In Situ Hybridization
Note:
For non-radioactive in situ Hybridization with alkaline
phosphatase and BM Purple AP Substrate it is important not to use xylene-based mounting media because
these could lead to crystal formations of the color pre
cipitates.
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Ordering Information
AP-Coupled Antibodies
Product
Anti-Digoxigenin-AP, 
Fab fragments, from sheep
Anti-Biotin AP, Fab fragments
Anti-Fluorescein-AP, Fab fragments
Streptavidin-AP
Pack size
150 U (200 ␮l)
Cat. No.
11 093 274 001
150 U
200 ␮l
150 U
150 U
1,000 U (1 ml)
11 426 303 001
11 426 338 001
11 093 266 001
11 089 161 001
Chromogene Substrate for Alkaline Phosphatases
Product
BCIP, crystals
BCIP, 4-Toluidinsalz, 
powder
BCIP, 4-toluidinesalt, solution
INT/BCIP Stock Solution
Fast Red Tablets
NBT/BCIP ready-to-use tablets
NBT/BCIP Stock Solution
NBT Solution
NBT crystals
Pack size
1g
250 mg
1g
3 ml
(150 mg)
3 ml
20 tablets
20 tablets
8 ml
3 ml
(300 mg)
1g
5g
Cat. No.
11 017 373 001
10 760 994 001
11 585 002 001
11 383 221 001
11 681 460 001
11 496 549 001
11 697 471 001
11 681 451 001
11 383 213 001
11 087 479 001
11 585 029 001
Labeling of Biomolecules
Product
DIG RNA Labeling Mix
DIG DNA Labeling Mix
DIG High Prime
Pack size
40 ␮l
(20 reactions)
50 ␮l
(25 reactions)
160 ␮l (40 labeling
reactions)
Cat. No.
11 277 073 910
11 277 065 910
11 585 606 910
Other reagents
Product
Tween 20, 10% (v/v)
Blocking Reagent
Trademark
Pack size
5 × 10 ml
50 g
Cat. No.
11 332 465 001
11 096 176 001
All brands or product names are trademarks of their
respective holders.
* available from Roche Applied Science
Contact and Support
Regulatory
Disclaimer
For life science research only. 
Not for use in diagnostic procedures.
To ask questions, solve problems, suggest enhancements or report new
applications, please visit our Online Technical Support Site at:
www.roche-applied-science.com/support
To call, write, fax, or email us, visit the Roche Applied Science home page,
www.roche-applied-science.com, and select your home country. Countryspecific contact information will be displayed. Use the Product Search function to find Pack Inserts and Material Safety Data Sheets.
Roche Diagnostics GmbH
Roche Applied Science
68298 Mannheim
Germany