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PBMC Culture advice

culture

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#1 allabouttheY

allabouttheY

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Posted 04 August 2014 - 10:20 AM

Hi all,

 

Some advice would be appreciated. The aim Storing Human cells at -80C using Mr. Frosty to control the rate of cellular freezing.

 

We had being originally using a PBMC protocol using 2 freezing medias one with RPMI and HSA, the second with HSA80% and DMSO 20%.

 

We are moving towards using BSA instead, however the region we are located at, reagents can at times be hard to obtain. The recommended Specifications for BSA is as followed

BSA specifications

  • Fatty acids free
  • Sterile
  • Azide/preservatives free
  • Aqueous solution                        

Desirable

  • Fraction V
  • Resolubilised crystalline factor V.

 

  • we don’t want a lyophilized powder form as to prevent additional sterility steps

 

The option available in our Area currently is

Bovine Serum Albumin solution
10% in DPBS, low endotoxin, fatty acid free, suitable for cell culture,
sterile-filtered

 

Firstly is this a good alternative from HSA?

Would the fact that it is in a 10% DPBS require a change in our HBBS cell washing part of the protocol?

 

RPMI does the ‘with or without’ Ca, Mg and phenol options have an effect on rate of cell recovery?

 

Suggestions as to a better protocol are also welcome.

 

Thank you

 







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