I would like to describe a problem I was faced with two days ago. For many of you this may be quite a simple problem but in the context that I manage a multidisciplinary laboratory that performs 3000 group and antibody screens and cross-matches 400 units per year we are quite small. I think it important for the smaller laboratories to share their problems, and this is my first thread.

A patient presented with a request for the provision of 2 units of blood for a spinal operation the following morning. Grouped as O Rh (D) negative with a positive antibody screen. Antibody I.D. showed that the patient had Anti-M. I should mention that we use Ortho BioVue technology, no automation. Thats the easy part dealt with.

The UK guidelines (I assume it is the same internationally) say that antigen negative blood should be selected if the Anti-M is reactive at 37°C otherwise blood selection by IAT crossmatch is acceptable. There in lies the problem, how do I know whether this Anti-M is reactive at 37°C or does it just react at RT. This was Friday afternoon and I pondered this for a little while before phoning the head of the local reference laboratory (Tooting blood centre) who just happens to be the ever helpful Malcom Needs. Malcolm and his team are used to dealing with multi-antibody problems but I can assure everyone out there they are just as helpful to the smaller laboratory. An important message for those who worry about asking simple questions.

Malcolm suggested we order historical M negative blood from the issues department. This was delivered to my lab in the late afternoon, cross-matched with the patient and was compatible and issued. Whilst waiting for the units to be delivered from Tooting we did cross-match the four O Rh(D) negative units we had in stock and found one to be compatible but not a confirmed M negative unit.

We always refer our samples for confirmation of antibody identity to the local reference laboratory (Malcolm Needs team), and I will be interested to know whether this Anti-M is reactive at 37°C. Malcolm did describe how they elucidate this, but I have I have deliberately left this out to encourage discussion unless you all know.

Steve Jeff

Steve,

I will not spoil your thread by making any comments s to how we do resolve the problems, but I would, nevertheless, like to thank you for your kind words, which I will pass on to my team.

Malcolm

:D:D

we do pick up anti-M in Gel which are IgM not IgG...we give crossmatch compatible(by Gel) units but do not type them for M.

Hi aakupaku

The question I ask, how do you know thay are IgM and not IgG antibodies. If they react at 37°C, then we have to give M negative red cells?

Steve:)

When we find an anti-M using gel we immediately switch to a less sensitive method -LISS. If you use PeG, you do not read after 37C incubation so that enhancement will not solve your anti-M problem. BY using LISS, we can determine if there is reactivity at 37C. We give crossmatch compatible in these instances, ie, if negative at 37 and AHG in LISS. I have seen M's that react with gel at AHG, but only the homozygous cells. I had a customer who sent an abid that was of this nature, they transfused 6 AHG compatible units and asked my to type those - I told them that I bet that all were M+. I was correct and the patient had no sequellae following those units.

Hi David

I am not an expert in transfusion serology. however, I query whether resorting to a less senistive method is the best scientific response. When I spoke to Malcolm Needs on Friday he did say they resorted to tube techniques probably in LISS, but more importantly incubation at RT and 37°C. The anti-M being particularly avid at RT.

What you have described may be the same process. I know malcolm will correct me if I have made the incorrect interpretation.

Steve:wave:

I'll hold my peace for now!

I have a written procedure for dealing with Anti-M that does detail that the testing be taken to tube when gel is positive. The only wayt to determine whether or not the M is warm reactive is to separate the phaes and any gel technology cannot accomplish that. I don't have an issue with reverting to a so called less sensitive technique at all. The gel is the first line and if it showed only M, then Tube is fine and full crossmatch will complete the picture.

I think what we do is fairly compliant with what Malcolm does . . . once again, as Lara says, if you are only dealing with anti-M then going less sensitive works fine.

Hi Lara

I think the procedure you describe in separating the phases using tubes and I assume pre-warming sera, LISS and cell suspensions is very much what Malcolm was describing to me.

In no way am I suggesting that a tube technique is less sensitive, in some eyes it remains the gold standard. I am sorry if I have confused the interpretation of the tube technique as being less senistive. I have to confess that I don't have access to tube technology anymore.

Steve

In addition to determining reactivity at 37, you can also treat the serum/plasma with 0.01 M DTT to destroy IgM reactivity and determine the presence of an IgG componet... I know many hospitals don't do this but reference labs usually can. (Note this procedure is mostly used for determining the class of Mama's Aby, at least here it is)

I read some where that most examples of anti-M do have an IgG component, and still they do not cause transfusion reactions when M positive units are transfused. We do not perform any special tests to determine if the reactiviity is only at the AHG phase or not. We perform AHG crossmatch for patient's with anti-M regardless of the phase of reactivity. No antigen typing required.

JB

It's always interesting to try to define "activity" or "reactivity" at 37o, since you can see in tube in the AHG phase:

1. "Carryover" agglutination in AHG, where IgM latches on in earlier phases and just doesn't let go during incubation and washing.

2. Complement bound, which few see since most of us use just anti-IgG.

3. IgG.

4. And what of the MM cells which react in AHG but the MN cells which don't - a blanket indictment of all M+ cells?

Since rare examples of IgM anti-M can bind complement at 37o, we have a conservative approach where we do prewarmed tube crossmatches using polyspecific AHG to cover all the bases and issue crossmatch compatible units. Even using poly, the prewarming eliminates the activity in AHG most of the time and it's not too hard to find XM-compatible units.

Since rare examples of IgM anti-M can bind complement at 37o, we have a conservative approach where we do prewarmed tube crossmatches using polyspecific AHG to cover all the bases and issue crossmatch compatible units. Even using poly, the prewarming eliminates the activity in AHG most of the time and it's not too hard to find XM-compatible units.

I've used this method at various places I've worked and it all seemed a lesson in futility because the predetermined result (negative) was almost always achieved. Of course, we'd do everything possible to be sure it was only an anti-M before prewarming (prewarming has created quite a controversy here in the Midwest.....).

Margaret W.

Hi

You may be interested to know that the reference report came back today confirming the presence of anti-M. Additionally the anti-M was not detected by LISS tube IAT at 37°C indicating that cross-match compatible blood would be acceptable for this patient. However, I can confirm Lara's statement above that gel/bead technology cannot be used to elucidate whether the antibody reacts at 37°C even if all the phases of the reactants are pre-warmed, because I tried.

I would still have to refer to the reference laboratory because I do not access to tube/LISS technology. However, the patient has been issued with an antibody card and is registered on the National Blood Transfusion service data base now.

Steve:wave:

OK, Anti-M's are kind of my forte, so to speak....

Anti-M's rarely read the text books and follow the rules on how they are supposed to react. Even several homozygous M cells on a panel might not react....

If you suspect Anti-M..but can't get it to "behave"...I suggest the following (in the Technical Manual)...

Increase your serum/plasma to cell ratio....(we normally use 50 m cells with 25 microliters of plasma)...but increasing your plasma amount to at least match your cell amount helps . Also incubation for 30 mins at RT will give you a lovely picture of an Anti-M. (If it's there).

If we detect anti-M in Gel, we issue gel crossmatch compatible units. For pregnant patient, we repeat tube screening(with PEG) and if reactive, perform prewarm...and if reative with prewarm we follow up with titer other wise we do not worry. For crossmatchin we always give gel crossmatch compatible RBC(do not give M- except neonates with mother has anti-M).

Issitt says that AHG xm compatible is sufficient for all patients with anti-M so that is what I have always done. Antigen negative not required. Have had to probe harder with pregnant moms, of course.