2 replies to this topic

[SF_HK]

Hi,

I'm new to IHC and would appreciate if my citrate buffer (ph6) could be verified.  I'm using this for antigen retrieval:

A) 0.1M Citric Acid
1.92 gm citric acid, anhydrous to 100ml distilled water


0.1M NaCitrate, dihydrate
14.7 gm NaCitrate dihydrate to 500ml distilled water



C) Working sloution: total 500ml
9.0 ml of 0.1 M Citric Acid (A)
41.0 ml of 0.1M NaCitrate, dihydrate (
add 250ml distilled water, pH to 6 with NaOH and bring final volume to 500ml using distilled water.

Thanks

[Clare]

Hello I can't remember actually making the citrate buffer ( it was AGES ago) but I know my buffer (10mM, pH 6) contained 0.05% Tween 20 and worked beautifully

Clare

PS:

Citrate Buffer (10mM Citric Acid, 0.05% Tween 20, pH 6.0):

      Citric acid (anhydrous)  --------------- 1.92 g

      Distilled water -------------------------- 1000 ml

      Mix to dissolve. Adjust pH to 6.0 with 1N NaOH and then add 0.5 ml of Tween 20 and mix well. Store this solution at room temperature for 3 months or at 4 C for longer storage.

SF_HK, on 16 September 2010 - 01:06 AM, said:

Hi,

I'm new to IHC and would appreciate if my citrate buffer (ph6) could be verified.  I'm using this for antigen retrieval:

A) 0.1M Citric Acid
1.92 gm citric acid, anhydrous to 100ml distilled water


0.1M NaCitrate, dihydrate
14.7 gm NaCitrate dihydrate to 500ml distilled water



C) Working sloution: total 500ml
9.0 ml of 0.1 M Citric Acid (A)
41.0 ml of 0.1M NaCitrate, dihydrate (
add 250ml distilled water, pH to 6 with NaOH and bring final volume to 500ml using distilled water.

Thanks

Edited by Clare, 16 September 2010 - 03:23 AM.

[mdfenko]

when preparing a buffer from equimolar solutions of an acid and base form of a buffering species, you adjust the pH by adding one or the other solution. this way you maintain the concentration of the buffering compound regardless of final volume. you can then prepare solutions by diluting this stock.

in your example, i would prepare a stock of the 0.1M citrate buffer, pH 6, without using naoh to adjust. then dilute 50 ml to 500.