Can anyone help me, I've been asked to prepare C3 and C4 sensitized cells, which I last prepared in 1982! Has anybody got an up to date method for this preparation?

Thanks Steve

We use fress serum and sucrose to sensitice cells with C4 (for detection of anti Ch/Rg antibodies), I know that these cells also have C3 on the surface but I have never tested it.

What do you want to do with those cells after sensitization?

Peter

It's to validate polyspecific cassettes for C3b and C3d

Steve

http://www.hemobioscience.com/PDFs/C3_Press_Release_Final.pdf

Have you seen this ad?

It are commercial C3 coated cells.

Yes I have thanks, but we'll need in house reagents.

Steve

I have made some copies from our Methodes book (from 1993) from the American Red Cross about C3b, C3d and C4 coating.

I hope this will be of use.

Peter

Thanks!

I have found that the "home made" complement cells have a very short life span.

Hi,

 

After prepping cells using the Fruitstone method (American Red Cross #139) the resulting cells are reacting with our Anti-C3 reagent AND our Monospecific Anti-IgG. I was expecting a positive response from the C3 reagent only. Does anyone have any idea of why IgG is non-specifically binding to the cells along with C3b?

 

Thx

Most methods for coating cells with complement will also coat them with a certain amount of IgG.  Whether you then detect the IgG depends a bit on the method and the reagent you are using.  Basically  most complement control cells are only suitable for checking that you have a positive reaction with anti-Complement reagents, but NOT for controlling that other reagents do not detect anti-complement

Thanks for the response. 

 

If this is the case, when we are controlling AHG tests ,which contain anti-IgG and anti-complement components, how can we be sure that any positive reaction is due to only complement coating of the cells and not IgG (or a combination of both)?

Question - why are you making these cells in-house when they are readily available from a manufacturer? Cost?

I am not based in a clinical lab, only require cells for research purposes. Commercial cells require at least a 12 month standing order, something i don't need. In any case i think i may have solved the problems i was having with non-specific IgG binding by altering my incubation procedure at 0-1C. Results now show i have pure C3b coating.