Tabbie

When I was working in RCI at NHSBT-Tooting Centre, we used to store our liquid reagent cells in Cellstab, but wash them an resuspend them in Dil2 for use.  We found the reactions we got were much, much sharper (I don't mean that the reactions were more sensitive to detecting weak antibodies, although we believed that they were, but that it was far easier to "see" the reactions as clear reactions, rather than "fuzzy" reactions), and, in addition, it meant that we didn't detect reactions caused by antibodies directed against the preservatives in the Cellstab.  We were able to show this with multiple photographs.
Despite all the evidence, we were told that we couled not continue to do this, as we were not standardised with the other NHSBT RCI Laboratories (standardisation is everything these days, even if it means dumbing down, rather than bringing everyone up to an excellent standard, and because it was more expensive.  The only problem was that we were able to show that it was actually LESS expensive, because it meant less testing, and no testing for antibodies against preservatives.
This did not fit with management theory, however, and so we had to stop.  Since then we got "fuzzy" reactions, leading to many cases of repeat testing, and many cases of antibodies against preservatives and, hence, more expensive testing in terms of reagents, staff time and fairly simple investigations into moderate or even complex investigations, but hey, what did we know!  At least we are now standardised (and expensive)!

Below is the exact text from the guidelines.    I agree that potentiators added to the reverse group may detect non ABO antibodies in addition to the ABO antibodies but who adds potentiators to reverse group routinely?   One reason that anti c is mentioned may be that reverse group cells are usually Rh neg.   I have seen strong IgG anti c react in reverse group with no potentiators. 
 
 
Other reverse grouping anomalies: Potentiators in the reverse grouping reagents may cause IgG antibodies such as anti-c to be detected in the reverse group.

I've seen anti-c reacting with the reverse grouping cells many times.  I've only seen anti-e reacting like that a very few times.

I just answered this question.


My Score PASS  

Way back in 2003, the AABB published "Guidelines for Implementing an Electronic Crossmatch". In the section REQUIREMENTS on page 4 "Additional criteria suggested, but not universally accepted, include: ... The antibody screen should include a three- or four-cell sample."
No further explanation was given but presumably it was for the reasons stated above that you are likely to have more cells with a homozygous antigen expression in a 3-cell panel than you are in a 2-cell panel. Most manufacturers of 3-cell panels make sure to include cells that are homozygous for Duffy and Kidd to avoid missing those.
So long story short....that's why we use a 3-cell panel as part of our due-diligence of providing blood by computer assisted crossmatch.

Ideally it is great to rule-out with at least one homozygous expression, but it is not always practical. The anti-K scenario is a prime example as discussed, but for those of you who insist on a homozygous cross-out: Do you really delay an identification of an anti-D and a potential transfusion, just to rule-out anti-E and anti-C?
Most hospitals do not have access to r'r' or r"r" cells, so would you insist that the sample be sent to a reference laboratory for resolution?
Ruling-out with heterozygous examples is an acceptable practice when the situation warrants it.  Obviously, one should look at the "big picture" and if you are seeing reactivity consistently with only a homozygous expression of a certain antigen, then you wouldn't want to ignore that and rule-out on only a heterozygous expression (or 3 for that matter).
My advice though: When doing a heterozygous cross-out make sure you are using a technique that is considered an enhancement for the antibody in question and if in doubt, either call your local reference laboratory for advice or ship it to them for resolution.

Personally, I think people are getting a bit "hung up" about this - not least the people who "write the rules", and I think that a lot of these problems are caused by the sloppy use of nomenclature (see my first ever post on BloodBankTalk back in March 2009 - before it became PathLabTalk).
When you are using antibody screening red cells, you are using, I presume, R1R1, R2R2 and possibly rr red cells, but are you actually?  These are "most likely genotypes", that are derived from phenotypes, but I very much doubt if the donors, from which these red cells have been derived, have been genotyped.  Even if they have been genotyped, it is extremely unlikely that full gene sequencing has taken place on the RHD and RHCE genes to determine, whether either the RHD gene is present in a homozygous state, a hemizygous state, or a heterozygous state, where one RHD gene is of the wild type and the other is mutated, and would produce a variant D antigen. or the RHCE gene is homozygous for the production of, in order, the C and e antigens, the c and E antigens and the c and e antigens, or whether they may be hemizygous with a silent gene (either with a normal RHD gene to encode a D-- or D.. haplotype, or with a silent RHD gene to encode an Rhnull haplotype), or whether they may be heterozygous with, for example, a normal C and e antigen and a CeS antigen in, for want of a better way of putting it (although this, strictly speaking, only applies to genes, rather than antigens) in trans.
The same principle applies to all of the other antigens and their genetic backgrounds.
So what am I saying?
I am saying that, day after day, we are excluding the presence of an anti-D, an anti-C and an anti-E (and, possibly, an anti-f) with screening cells that we actually do not know what antigens they are actually expressing, and yet we do not worry about this.  It is only when we detect a reaction with the screening red cells that we bother to perform an antibody investigation to determine the specificity of the antibody causing the reaction.  What, I ask, does this say about our logic?  I would contend it says nothing flattering!  So, if you are detecting an anti-D, and you (that is the collective "you", JustaKIDD, not a personal "you" - I am certainly NOT attacking you as an individual) are worried about the presence of an anti-C or an anti-E, because you only have one example of r'r or r"r red cells to exclude the specificities, I, unlike the people who "write the rules" would say "so what", but, if you are worried, why not just assume the antibody/antibodies is/are present, and cross-match rr units (if, indeed, the red cells in the bag actually ARE rr ).
Lastly, in all the years since anti-C and anti-E have been described (1941 and 1943 respectively), how many patients, with anti-D in their plasma, have had a fatal transfusion reaction due to the presence of an undetected anti-C and/or anti-E?  Not many (if any) I would suggest, and yet we, as a whole, get ""hung up" about such things, for no apparent reasons.

No, what I meant was that certain antibodies (some examples of anti-H from a true Oh individual, most examples of anti-Vel, most examples anti-PPkP1 from pp individuals and some examples of anti-I from ii adults, together with some examples of anti-Jka and anti-Lea, plus, of course, some examples of anti-A, anti-B and anti-A,B) would cause antigen positive red cells in an antibody identification test to haemolyse, IF serum from a clotted sample was used (hence, in vitro tests), but if EDTA plasma was used, you would not see this haemolysis.  In the good old days, seeing haemolysis with all the red cells of an antibody panel, when using serum from a clotted sample, gave us a clue that we were dealing with one of the above antibody specificities (except, of course, the ABO antibodies - because the panel cells were all group O).  Nowadays, with the samples almost always being EDTA, we no longer have that clue!

Sorry Tabbie, but low prevalence antigens are antigens that are only expressed on the red cells of a few individuals in a particular population, or even worldwide, whereas low affinity antigens are antigens that may be expressed on any number of individual's red cells, but which do not "fit" well with their cognate antibodies.  In other words, there is only a weak link between the antigen and antibody, and these links can also easily be broken.  This is because antibody/antigen reactions follow The Law of Mass Action.  In the case of a low affinity antigen, the reverse reaction (the dissociation between the antigen and antibody) is at least as quick and easy as is the forward reaction (the association between the antibody and the antigen).  In this case, YES, repeated washing with saline can wash away the antibody, and YES, the eluate may be unreactive, but this is very rare.  That notwithstanding, the commercial companies have to write this into their inserts, to cover themselves in law.

If you haven't already, you should keep a copy of this response, Malcolm.  I have lost track of how many times this issue has come up here!
Scott

CD = chloroquine diphosphate. A chemical treatment to remove immunoglobulins from red cells, in the hope of getting a negative DAT, thereby allowing the use of antiglobulin-reactive antisera without interference from a positive DAT.

Never say never - bet you get an Rhnull next week!!!!!!!!!!!!!!!!!!!!!!!!

A classic case of don't believe EVERYTHING you read!  

It is the latter yan xia.
The various metal ions have no effect on either sensitisation or agglutination, but, are vital co-factors for the complete initiation of the classical complement pathway (C1qrs), leading to the membrane attack complex (MAC), and it is this MAC that allows haemolysis to take place, and it is this haemolysis that allows the Hb to escape from the red cells, which we see as a pink/red colour in our tests, which equates to a positive result.  As the classical complement pathway is a huge amplification system (one C1qrs complex, for example, results in the generation of some 8, 000 C3b molecules), it makes the test very sensitive indeed - much more so than just looking for agglutination.

An autocontrol is an indirect antiglobulin test that includes both patient serum/plasma and patient rbc incubated together at 37C and subsequently tested with antihuman globulin reagent.  Methodology includes the standard tube test, Gel test, PEG test etc.

When we automated (Echo), we stopped doing autocontrols routinely with panels. It's not even an available test on the Echo. If all screen or panel cells are positive that's a different story. We can run a DAT or crossmatch the patient with their own cells to give us an autocontrol. If we need to move to tube testing to resolve a problem I require the auto control using whatever enhancement media is being used for the panel/screen. If the auto is positive, we do a DAT. If there is suspicion of a hemolytic transfusion reaction or a hemolytic process we would do an auto and/or a DAT. Obviously a reference lab is going to approach things in a different way than a transfusion service.
I think that the sensitivity of the method, gel or solid phase, can be a bit of a curse - you find things that are not meaningful for transfusion purposes by doing autos on everyone. If the antibody is straight forward and there is no evidence or history of hemolysis/WAIHA, does the positive auto give us information of value? We've been doing it this way for almost 8 years now and it hasn't caused any issues, not even once. That positive autocontrol just gives you hornets to chase most of the time and doesn't change how you transfuse the patient. As my boss likes to ask - "Does it add value to the process?" If it doesn't, don't do it.

Thanks Malcolm.
(Actually, I was a bit confused about the whole McLeod nomenclature thing, the rest was just to get your attention.)
Scott

Sorry Scott, but an awful lot of this is wrong!  Certainly K and k (NOT K1 and K2, as explained above) were named after two pregnant women, and anti-K had primacy over anti-k, but anti-k (as mentioned in both The Blood Group Antigen FactsBook AND Bloody Brilliant!  A History of Blood Groups and Blood Groupers.) the woman involved in the discovery of anti-k was actually named Nocella, rather than Cellano.  Anti-K was described in 1946 in  the paper by Coombs, Mourant and Race that described the Direct Antiglobulin Technique, whereas anti-k was not described until 1949 in a paper by Levine, Backer, Wigod and Ponder.  All of that having been said, the actual first antibody/antigen that was described that is now in the Kell Blood Group System was Levay (now Kpc or KEL21), which was described by Callender, Race and Paykoc in 1945, but, of course, it was not recognised as part of any System at the time, as, for years and years, there was only one known example of anti-Levay, otherwise the Kell Blood Group System would have been called the Levay Blood Group System, and not the Cellano Blood Group System.
Mr Hugh McLeod, a dental student, was not a true Ko (or Kellnull), but had weak expression of Kell Blood Group System antigens.  True, his phenotype was described in 1961 (by Allen, Krabbe and Corcoran), but the true Ko (or Kellnull) phenotype, which, unlike the McLeod phenotype, has no expression of the Kell antigens whatsoever, and which was first described in 1957, just after Allen and Lewis had described anti-Kpa.  Such individuals usually produce an anti-Ku (the "u" standing for universal).
The McLeod phenotype and the McLeod Syndrome are by no mean synonymous.  Indeed, there are three interwoven conditions (for want of a better way of putting it).  There is the McLeod phenotype, as described above, which results in weaken expression of the antigens of the Kell Blood Group System, but such individuals are physically and mentally well.  Then there are individuals who have X-linked Chronic Granulomatous Disease (or CGD), but do not have the McLeod phenotype.  Thirdly, there are individuals who have both the McLeod phenotype and CGD, a condition that is known as the McLeod Syndrome (which is a bit strange, because the first person to be described who had the McLeod phenotype and CGD was actually named Claas).  Both the XK and the X-linked form of CGD are mapped to Xp21.1.
In 2010, a paper was published that suggested that King Henry VIII (not his father King Henry Vii) had McLeod Syndrome (Banks-Whitley C, Kramer K.  A new explanation for the reproductive woes and midlife decline of Henry VIII.  The Histoical Journal 2010; 53 (4): 827-848).  Three years later, A similar paper was published (Stride P, Lopes-Floro K.  Henry VIII, McLeod syndrome and Jacquetta's curse.  J R Coll Physicians Edin 2013; 43: 353-360).  These two papers are perfect examples of a little knowledge being a dangerous thing!!!!!!!!!!!!!
I will be talking about the Kell and Kx Blood Group Systems, and about Henry VIII and CGD in Providence, Rhode Island in May.

Annan, please see this quote from above.
K1 is incorrect (as quoted from The Blood Group Antigen FactsBook) and there is no such antigen as Kell.

Sorry John, but you are incorrect - and correct!  The first antigen within the Kell Blood Group System is, under the ISBT numerical system 006 001 (006 being the number of the Blood Group System, and 001 being the first antigen within that system) and, indeed, the 006 is never used in terms of Blood Group Systems (except on computers), and the "00" bit of the "001" is redundant, so you would think that K is equivalent to K1, but it isn't!  To quote from Reid ME, Lomas-Francis C, Olsson ML.  The Blood Group Antigen FactsBook."  3rd edition, 2012, Academic Press, page 3-7, "It is incorrect to refer to the K and k antigens as, respectively, K1 and K2; in the numerical terminology they should be referred to as KEL1 and KEL2."

I have been biting my tongue, trying not to say anything, but I have just got to!
Kell is the name of a Blood Group System, but the first antigen within the system is named K, and the antibody against it is named anti-K.
Those of you who screen for "Kell" and find negative donations are finding an awful lot of Ko donors (whereas the rest of the world is trying desperately to find some to freeze down), and those of you who are finding all these examples of "anti-Kell" are finding an awful lot of examples of anti-Ku, and in 43 years working in Reference Laboratories, I have only seen one example!

I've ony screened for Kell when the patient was sensitized.  If I had a large sickle population I would screen for it there also.  The newest screening we did was for folks on anti-CD38 therapy if we were unable to get a K typing prior to initiation or if they were K negative.

We always found one or two patients with a new anti-K every year.  The good thing is that K= blood is easy to find.  Oddly enough my wife, a nurse of course, has an anti-K which was one of the first antibodies I identified while still in school.  Luckily I am K= so that was never an issue with our children.  Her anti-D on the other was a much more significant bother.  Her anti-S has not been an issue either.  Some how I suspect she is what we fondly refer to as a "responder".   
 

The K antigen is the most immunogenic of the Kell Blood Group System antigens because the Thr193Met mutation voids an N-glycosylation site (asparegine 191), exposing a highly immunogenic area devoid of an N-linked sugar, and so I would be amazed if there had not (Lee S, Wu X, Reid M, Zelinski T, Redman C.  Molecular basis of the Kell (K1) phenotype.  Blood 1995; 85: 912-916.).

No problem - but there are reasons for this.
Although Rh antibody/antigen reactions can show dosage, it is comparatively rare for them to so do.  In addition, say we are talking about anti-C, the most common antibody specificity found with anti-C is, of course, anti-D, and so we (NHSBT) would have to provide hospitals with, not only r'r' red cells in the antibody panel, but also r'r' red cells for the screening cells - and we couldn't; they are far too rare.
The same applies to antigens within the Kell Blood Group System.  Although we (NHSBT) are able to provide an example of K+k- red cells in the antibody panel, we could not provide one in the screening cells; there are just not enough available.
In a case where a patient has already produced a clinically significant antibody, such as anti-Fya, BCSH Guidelines suggest that further blood transfusions should not only be Fy(a-), but also K-.  Personally, I think that we should be duty bound to ensure that the patient is not K+k-, so that they do not make an anti-k, but that is what the Guidelines say (and if the patient is K+k+, it doesn't matter anyway).