SILVER STAINING

[Dr. David Martin]

In article <595too$12b at mserv1.dl.ac.uk>,
   Dr Alok Adholaya <aloka at teri.ernet.in> wrote:
+>Dear Netters,
+>
+>I am in a look out for a protocol for silver staining of proteins
+>seperated on SDS-PAGE. The protocol should  have sensitivity of 
+>conventional silver staining  yet be rapid ( should not take more
+>than 15 minutes ).  Does anyone know of such a procedure ??????
+>

The extremely rapid silver stain follows: For best results use 
spotlessly clean glass dishes/plates and do not touch the gel at all.
 
It is not as good a stain as taking several hours to do it properly 
(background can be high) but it is sensitive and it does work.

.d

Method: Very Rapid Silver Stain

Chemicals required:	Coomassie Destain (40% MeOH 10% AcOH)
				5 mg/ml Dithiothreitol
				1 mg/ml AgNO3 10X
				15% Na2CO3
				40% Formaldehyde
				2.3M Citric Acid

Health and Safety:	40% Formaldehyde. Toxic in contact with skin. 
Wear gloves.

Procedure:	

	1. Microwave gel for 1 minute at full power in 100 ml 
Coomassie Destain.

	2. Wash briefly in dH2O to re hydrate the gel (5-10 mins or 
1min in microwave).

	3. Microwave gel for 1 minute at full power in 100 ml dH2O 
containing 100ul 5mg/ml DTT. 

	4. Wash in 0.1 mg/ml AgNO3 for 45 minutes to overnight.

	5. Wash briefly in dH2O.

	6. Wash in developer (40 ml 15% Na2CO3 200 ul 40% Formaldehyde 
in 200 ml) till solution goes brown (100 ml for approx 1 minute). 

	7. Changge developer and develop. Stop by adding 5ml 2.3M 
citric acid.

* David Martin - Atherosclerosis and Thrombosis research group  *
* http://www.uio.no/~damartin/      david.martin at biotek.uio.no  *
* Lab +47 22 95 84 54 Fax +47 22 69 41 30  GSM +47 90 74 27 65  *