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Author Topic: Wash buffer for the IHC  (Read 7084 times)

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Offline XueYuan

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Wash buffer for the IHC
« on: August 29, 2012, 02:10:11 PM »
Hi, I was use the abcam IHC-P protocol for my IHC and I found there is no 10XTBS receipt in that protocol. I called the company and they told me any receipt of TBS is worked in the IHC. So I looked on line and found that in the IHC world there are the receipt(http://www.ihcworld.com/_protocols/washing_buffers/tbs.htm). But  I wonder why the PH here is 8.4. I also found some other TBS receipt which PH is between 7.4-7.6. Does anybody know why the PH need to be high as 8.4? Also what is the best TBS for the IHC? Thank you!

Wash buffer for the IHC
« on: August 29, 2012, 02:10:11 PM »

Offline BJessica2

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Re: Wash buffer for the IHC
« Reply #1 on: September 10, 2012, 10:23:59 AM »
Hello,
The recipe below is a good one for TBS! By receipt you mean, RECIPE!!  :D this is a 10x solution that you can use at 1x.

TBS 10x (concentrated Tris-Buffered Saline), 1 liter   
24 g Tris base (Formula weight: 121.1 g)
88 g NaCl (Formula weight: 58.4 g)
Dissolve in 900 ml distilled water.                     
pH to 7.6 with 12N HCl.                        
Add distilled water to a final volume of 1 liter.                                                   
For a 1X solution, mix 1 part of the 10X solution with 9 parts distilled water and pH to 7.6 again.   
The final molar concentrations of the 1X solution are 20 mM Tris and 150 mM NaCl.                                    
An alternative recipe for Tris buffer combines Tris base and Tris-HCl. This avoids the large volume of potentially hazardous hydrochloric acid that is needed to neutralize a solution of Tris base alone.                                                   
The pH of the solution should be about 7.6 at room temperature. If too basic, adjust to pH 7.6 with concentrated HCl, and if too acidic, adjust with concentrated NaOH.                                          
Add distilled water to a final volume of 1 liter.                                                   

Offline XueYuan

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Re: Wash buffer for the IHC
« Reply #2 on: September 17, 2012, 01:57:21 PM »
Thank you for pointing out the mistake. :-[  And also thank you for sharing the TBS recipe.

Offline MT Scientist

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Re: Wash buffer for the IHC
« Reply #3 on: September 19, 2012, 04:42:39 PM »
As an aside - Receipt and recipe mean the same in this case.  "Receipt" is not currently/commonly used in the USA, though it appears to have been more common in the 18th and 19th century.

Re: Wash buffer for the IHC
« Reply #3 on: September 19, 2012, 04:42:39 PM »