Malcolm Needs

Hi Bianca, I agree that Anna's response was brilliant. DO NOT WORRY about the anti-Cw (or anything else come to that). As you have never had a transfusion and this is your first pregnancy, the chances of the anti-Cw being clinically significant to either you or your unborn baby are as near nil as could ever be. Your hospital is "covered" by the NHSBT-Tooting Centre and, if your hospital thinks it worthwhile, we will monitor the antibody during your pregnancy, but it depends on the titre (strength of the antibody) in your plasma. I am prepared to be my mortgage (even after yesterday's budget!) that your anti-Cw is extremely weak, and we would not even bother monitoring it after 28 weeks gestation, and would certainly NOT advocate testing your partner's blood. If you can convince your obstetrician that this should be done (and, if would give you total peace of mind, go for it) we would gladly perform the test, but it really isn't a worry. IF, AS YOU SAY, YOU HAVE NEVER BEEN TRANSFUSED OR BEEN PREGNANT BEFORE, THIS ANTIBODY WILL CAUSE NO PROBLEMS WHATSOEVER. Indeed, most cells used to screen for antibodies in the UK are NOT Cw+, so confident are we that the antibody is clinically insignificant in cases such as yours.

Hi David, Yes, I can see what you mean. We've had a couple of cases involving anti-Era in the last couple of years and, as far as I know, there are NO licensed anti-Era grouping reagents in the world. We had to use some SCARF derived antisera to group the two patients, using positive and negative controls, as there was no alternative. Proving that there were no other underlying atypical alloantibodies was fun! Fortunately, we had 6 other examples of ABO compatible Er(a-) cells available to us, and were able to rule out everything apart from a potential anti-K using these and the patient's own typing, and giving K- blood was no problem.

I agree with both of these posts, but, as David says, validation is an absolute MUST. We do almost all of our typing in gels now, with very few now performed in tubes.

Yes, I'd like to know that too!

I agree entirely with most of the authors in this thread. I am the manager of the red cell immunohaematology department at NHSBT-Tooting Centre in London and, whilst we use gel as a "first defence" in all cases, we would be totally lost without tube technique, particularly in cases of autoimmune haemolytic anaemia, and in particular within that group, cold autoimmune haemolytic anaemia, where pre-warming of the reactants is vital.

There are two things to remember about this. The first is that "cold" reacting antibodies can sensitize red cells extremely quickly as the tests are set up. The gel tests are usually set up at room temperature and then moved to a 37oC environment for incubation, and then the centrifugation at the end of incubation is performed, once again, at room temperature. Therefore, "cold" reacting anti-M's (and "cold" reacting antibodies of other specificities) have plenty of opportunity to sensitize (and, therefore, agglutinate) red cells during the gel IAT, but, this is a kind of "false positive", as such antibodies are not truly "warm" reacting. The second thing is that, to a certain extent, the immunoglobulin class of the antibody is irrelevant, as some IgG antibodies react optimally in the cold (for example, the anti-P in PCH - the antibody itself is not what causes the problem here - it is the overlap of the temperature at which the antibody reacts and the temperature at which complement is activated - hence the bi-phasic temperature test). My own, very strongly held view is that, unless an anti-M reacts by the tube IAT technique, performed strictly at 37oC, we pre-warmed plasma and red cell suspension prior to mixing, and washing with pre-warmed saline prior to the addition of the AHG, it is not clinically significant, and cross-match compatible blood is more than suitable.

One of the ways around this is the use of Radio Frequency Identification (RFID). This is by no means inexpensive, may not be idiot proof, but is as near foolproof as I have seen. The sample label is produced at the point of phlebotomy with a hand held device by scanning a bar code on the patient's wrist band. The label is produced with eye readable patient identification and a copy of the patient's bar code. This barcode can be read by the automation in the Blood Bank and there is, therefore, no requirement to relabel. Pre-printed "addressograph" labels, as I think are described by JALOMAHE, are more or less "banned" in the UK by the British Council for Standards in Haematology (BCSH) Guidelines, whilst RFID labels produced at the point of phlebotomy are positively encouraged - but they are expensive. I am certain, as has happened before, they will become cheaper in a very few years. After all, I can remember when abacusses were expensive, let alone, calculators and computers! One thing that does worry me is that you say, JALOMAHE, that this is the first such occasion in 20 years. This may be the first time that such a situation has resulted in clinical sequalae, but do you know how many times it has happened when, for example, two group O, D+ patients' samples have been swapped, resulting in a "safe" (and I use the term in its loosest sense) transfusion?

Galvania and Eoin are quite correct in my opinion. If you do not use such a negative control, but use things like saline, albumin, saline and albumin, etc, you are not using a negative control; you are using a sop (and I don't mean a Standard Operating Procedure!) to make you think you are using a negative control. It gives you peace of mind when, in reality, you should be extremely nervous!

I couldn't agree more with Ellen Zeigler. The use of controls, particularly negative controls, with such antisera as an anti-U, whilst just screening, is a criminal waste of a very precious reagent cell.

I cannot see any problem with this for screening purposes, but I would have huge problems with the idea if the outdated antisera were used to actually type patients or any units of blood destined to be transfused to a patient with anti-Kpa, anti-k, anti-U, etc, unless they were then fully checked with a licensed reagent. I am fully aware that these reagents can be/are expensive, but I also think that, if they are available, and something went wrong, and they had not been used, there could be big trouble!

Hi Johnny, I replied to you, but it has come out on "Screening Antigen Negative Units???" for some reason! Malcolm:confused:

This, of course, is of no relevance to people in the USA, but in the UK we are "governed" by Guidelines issued by the British Council for Standards in Haematology. In respect of antibody identification, these state that to positively identify the specificity of an antibody, the plasma must react with two examples of red cells expressing the antigen (usually as a "double dose"), and not react with two examples of red cells that lack expression of the antigen. In addition, antibody specificities against other clinically significant atypical alloantibodies directed against the major blood group antigens must be excluded. This can be exceedingly difficult on occasions (e.g. when the antibody is directed against a very high frequency antigen, such as Rh29, or when there is a complex mixture of "common" antibodies). In such cases, it is worth remembering the usefulness of adsorption and elution to elucidate the case. One word of caution. If you are faced with identifying the specificity of an antibody against a low frequency antigen (although this exercise is usually esoteric, rather than useful!) be aware that many red cells labelled as positive for a particular low frequency antigen may, in fact, NOT be positive for this antigen. Unless they have been typed using antisera that are known to be monospecific, or have been checked at a molecular level, they may have reacted with a second specificity within the grouping reagent, as individuals making an antibody directed against a low frequency antigen often produce a "soup" of antibody specificities against low frequency antigens.

This is quite true, but the enzymes used are very different from the enzyme found in the green coffee bean used to convert group B red cells to group "O" red cells some years ago. The enzymes used now work at a pH nearer to physiological pH and are more efficient. Whereas the enzymes used earlier were unlikely ever to be used at an "industrial level", the enzymes used by Martin Olsson and his group in Sweden seem capable of this upgrade and they have found an enzyme that will convert A2 red cells to group "O".

I agree wholeheartedly about the usefulness of the FactsBook. I am the Reference Service Manager of Red Cell Immunohaematology at the NHSBT-Tooting Centre in London, and we use this book almost daily (in fact, it is beginning to fall apart we use it so much). It is quite brilliant. For those who would require a more basic book, I would thoroughly recommend Essential Guide to Blood Groups, by Geoff Daniels and Imelda Bromilow, Blackwell Publishing 2007 (ISBN-13: 978-1-4051-5349-2 and ISBN-10: 1-4051-5349-0). I believe it is published by Blackwell Publishing inc, Massachusetts in the USA. I'm not sure of the cost (particularly in dollars) but it is not expensive.

I entirely agree that only anti-IgG is required for testing cord blood. That having been said, I have seen several examples of maternal Kidd antibodies causing a positive DAT with both anti-IgG and anti-C3d, albeit weakly with the anti-C3d. I have never seen a case of clinically significant HDN/HDF caused by a Kidd antibody, although I am well aware of isolated cases being in the literature.

Rania, if I could have an email address, I've just completed an essay on this subject for my Continuing Professional Development file. Whilst I would not say that what I have written is by any means a brilliant piece of scientific work (actually, far from it), the references I cite may well be of use to you (but I really would suggest that you go back to the original references than rely on my own ramblings)!

Yes, that is what I meant by FYA/FY. The phenotype Fy(a+b-) may actually be heterozygous expression of the Fy(a) antigen (and I know that I've used bad terminology - one cannot have heterozygosity of an antigen, only a gene!).

I'm just wondering why rcurrie would give blood that is Duffy matched? When you consider that it is likely that about 68% of your sickle population will group as Fy(a-b-), but the vast majority of these will have an FYB gene (granted, not expressed on the red cells), it would seem a bit of a waste of Fy(a-b-) blood when this may be required, sometimes in large amounts, for a sickle cell patient with a genuine FY/FY genotype and anti-Fy3. It could be, of course, something as simple as that you have more Fy(a-b-) donors than we do in Britain, and that blood is more available to you.

Whatever method you use, make sure that your cells are fresh, as the Fy(a) and Fy( antigens decrease in strength with storage, and, if you possibly can (not easy, I know) ensure that your cells are FYA/FYA and not FYA/FY. Remember, anti-Fya will often show "dosage".

Rashmi, with a bit of luck, the two Powerpoint lectures I sent you should help!

I could not agree more with Marilyn Moulds. Until the specificity is properly determined, calling anything "HTLA" (I don't like the term either!) is very dangerous.

I'm afraid that I have come to the party a bit late, but that having been said, I think that it may well be worthwhile saying the following. In the Laboratory at NHSBT-Tooting Centre, London, where I am the Reference Service Manager, we have run a screening programme to detect rare donors for well over 25 years. We use, mainly, human derived antisera for this, although we now have access to some monoclonal antibodies directed against high incidence antigens. During this time, we have screened, quite literally hundreds of thousands of donors for the presence or, more interestingly, the absence of such antigens as U, Lub, k, Kpb, Jsb, Coa, I, Vel, Lan, P+P1+Pk and Yta (to name but a few. For the last 10 years, we have kept accurate figures, and have performed 1, 514, 436 tests and found 2, 025 rare donors as a consequence (including 724 K+k- donors, 32 Kp(a+b-) donors, 86 Lu(a-b-) donors, 54 Co(a-b+) donors, 51 Vel- donors and 19 Yt(a-b+) donors). In order to use the human derived antisera efficiently, usually obtained from a full donation from an "ex-patient", we perform titrations to find the end point, take the dilution back a couple of steps, and then use this dilution to screen (for example, we use our human derived anti-Lan at a dilution of 1 in 100, and have discovered 9 Lan- donors during this time). We do use positive and negative controls, but, in our case, I'm not sure why we need the positive control, because if the antisera had stopped working, we would soon know! Every donor would be Vel-! After we have done our screening, we then either test the donor's blood with accredited antisera "in house", or send a sample to the International Blood Group Reference Laboratory at NHSBT-Filton Centre, Bristol, for confirmation. Our screening programme seems to me just to be somewhat larger version of what has been suggested by Skinrash, and I would agree with most of the answers posted that there is nothing wrong with this approach. It is cheap and it is effective. I also agree wholeheartedly with those that have said that you do not become a manufacturer. You are checking your results with accredeted antisera after your screen. You are not relying on the original results. I say good luck to you, and there should be encouragement for this kind of enterprise.

The answer is that you can't, but you can make a very educated guess. The fact that the agglutination disappears once the tests are incubated strongly suggests the presence of a "cold" reacting antibody, and the fact that the DAT reacts only with anti-C3 also suggests a "cold" antibody. There is a very strong chance, therefore, that the antibody is auto-anti-H or auto-anti-HI, but without performing an auto control (which takes very little time and will not be that expensive) an auto-antibody cannot be proved. That having been said, the chances of the antibody being clinically significant (in terms of a transfusion) are disappearingly small.