Following the manufacturer’s IFU (Ortho; tube method), when performing antigen typing for CEce, we test IS, followed by RT incubation, mix, centrifuge, grade.

For Jka/b, Lea/b, K and P1 (procedure is the same, except these have no IS read prior to incubation), we incubate, do not mix (as mixing after incubation is not mentioned in the IFU), centrifuge, grade.

So, per manufacturer’s instructions, we mix before and after RT incubation for CEce testing but not for the other direct agglutination reagents, where we only mix before incubation at RT and spin without mixing.

Does this not mess with anyone else’s head? I understand that sensitization occurs during incubation and that any mixing after an incubation is futile (I think I’m correct on that?).

Thus,

Question 1: What would be the point of mixing our tubes after incubation in any Rh antigen typing procedure?

Question 2: I am under the impression that unnecessary mixing can break up weak hemagglutination, so wouldn’t post-incubation mixing carry with it the distinct possibility of false-negative reactions (yes, I know MOST donors react strongly with CE/ce)?

It has always been an unspoken rule for me to gently mix it once before every centrifugation. For the Jka/b, you mixed it once then incubate and then centrifuge. 

For the CEce you mixed it once for the IS before centrifuge. Then for the RT incubation part, you mixed it once before centrifuge. 

The gentle mixing before centrifugation allows cells to resuspend and bind to any possible "unbound" antibodies and then latticed together in the centrifuge. 

I attached the Ortho Instructions for Use for Anti-CEce. Good luck. 

Ortho_CcEe_.pdf