Malcolm Needs

Thank you for your kind comment David.
 
Anti-M is the bane of my life!
 
Until relatively recently, if an anti-M was referred to my laboratory, we would test it first by Bio-Rad (Used to be Dia-Med) gel IAT and enzyme technique (enzyme technique to see if anything else is there - even I know that the M antigen is destroyed by papain!!!!!!!!!) to make sure that there was an anti-M present.  If there was an anti-M present, we would then test it by pre-warmed, warm-washed LISS tube IAT at 37oC.  If we saw no reactions by this technique, we would advise that cross-match compatible blood would be suitable, and safe, for transfusion.  If we did see reactions by this technique, we would advise that M- typed blood should be cross-matched, and those compatible transfused.
 
The reason for the difference between the gel technique and the tube technique is two-fold.
 
Firstly, in the gel technique, the reactants (plasma and red cells) are introduced to each other at room temperature, and then the cassettes are put into a 37oC incubator.  "Cold-reactive" antibodies can sensitise red cells in "peco-seconds" (I may have exaggerated a bit there!!!!!!!!!!!), but the antibody-antigen complex takes a bit of time to dissociate (longer than the incubation time), and so a "false IAT positive reaction" is seen.  On the other hand, if the reactants are introduced to each other, already at 37oC, "cold-reacting" anti-M will (eventually) sensitise red cells, but not in the time allowed for incubation, and so the reaction would be negative.
 
The second reason is that the column containing the AHG is slightly acidic, and if there is one antibody/antigen reaction that likes acidic reactions, it is that between anti-M and the M antigen.
 
I was quite happy with this situation, BUT, and my own line manager is well aware of this, so I don't mind saying it publically, NHSBT has decided, on the grounds that most of our hospitals use gel techniques, we now recommend giving M- typed blood to anyone who has an anti-M, which I think is a total waste of units of blood that have been typed for all sorts of other antigens and have been found to have (presumed) homozygous expression for other, more useful, antigens.
 
I am well aware of the fact that anti-M can cause haemolytic transfusion reactions (IF serologically reactive at 37oC) and can cause clinically significant haemolytic disease of the foetus and newborn (IF serologically reactive at 37oC), and, in the case of HDFN, particularly amongst ethnicities in the Far East, such as the Japanese, irrespective of titre, but these cases are very, very rare.

Thank you for your kind comment David.
 
Anti-M is the bane of my life!
 
Until relatively recently, if an anti-M was referred to my laboratory, we would test it first by Bio-Rad (Used to be Dia-Med) gel IAT and enzyme technique (enzyme technique to see if anything else is there - even I know that the M antigen is destroyed by papain!!!!!!!!!) to make sure that there was an anti-M present.  If there was an anti-M present, we would then test it by pre-warmed, warm-washed LISS tube IAT at 37oC.  If we saw no reactions by this technique, we would advise that cross-match compatible blood would be suitable, and safe, for transfusion.  If we did see reactions by this technique, we would advise that M- typed blood should be cross-matched, and those compatible transfused.
 
The reason for the difference between the gel technique and the tube technique is two-fold.
 
Firstly, in the gel technique, the reactants (plasma and red cells) are introduced to each other at room temperature, and then the cassettes are put into a 37oC incubator.  "Cold-reactive" antibodies can sensitise red cells in "peco-seconds" (I may have exaggerated a bit there!!!!!!!!!!!), but the antibody-antigen complex takes a bit of time to dissociate (longer than the incubation time), and so a "false IAT positive reaction" is seen.  On the other hand, if the reactants are introduced to each other, already at 37oC, "cold-reacting" anti-M will (eventually) sensitise red cells, but not in the time allowed for incubation, and so the reaction would be negative.
 
The second reason is that the column containing the AHG is slightly acidic, and if there is one antibody/antigen reaction that likes acidic reactions, it is that between anti-M and the M antigen.
 
I was quite happy with this situation, BUT, and my own line manager is well aware of this, so I don't mind saying it publically, NHSBT has decided, on the grounds that most of our hospitals use gel techniques, we now recommend giving M- typed blood to anyone who has an anti-M, which I think is a total waste of units of blood that have been typed for all sorts of other antigens and have been found to have (presumed) homozygous expression for other, more useful, antigens.
 
I am well aware of the fact that anti-M can cause haemolytic transfusion reactions (IF serologically reactive at 37oC) and can cause clinically significant haemolytic disease of the foetus and newborn (IF serologically reactive at 37oC), and, in the case of HDFN, particularly amongst ethnicities in the Far East, such as the Japanese, irrespective of titre, but these cases are very, very rare.

Testing for antiG.ppt
With a bit of luck, this may explain a bit easier what I was trying to explain in words.

Hi Sara,
The first clue that the antibody may be anti-G, or anti-C+G, rather than anti-C+D, is indeed that the titre of the anti-C is higher than that of the anti-D, but the fact that the anti-C titre is higher than that of the anti-D is only a clue. It could be that you actually have an anti-C+D where, coincidentally, the anti-C titre is higher than the anti-D. It is absolutely essential, therefore, that you can prove that no anti-D is present.
One way of doing this is to divide the patient's plasma sample into two.
The first sample is adsorbed using Ro red cells treated with a proteolytic enzyme, such as papain or ficin, which would adsorb out any anti-G and any anti-D present, but would leave any anti-C present. At the end of the adsorption process (about 4 cycles), this plasma is tested against r'r red cells by IAT. If there is a positive reaction, then the original plasma contained anti-C and, possibly, anti-G. If there is a negative reaction, then the original plasma contained, possibly, anti-G and anti-D, but not anti-C.
The second sample is adsorbed using r'r red cells treated with a proteolytic enzyme, such as papain or ficin, which would adsorb out any anti-G and any anti-C present, but would leave any anti-D present. At the end of the adsorption process (about 4 cycles), this plasma is tested against Ro red cells by IAT. If there is a positive reaction, then the original plasma contained anti-D and, possibly, anti-G. If there is a negative reaction, then the original plasma contained, possibly, anti-G and anti-C, but not anti-D.
So, if there are no reactions with the plasma adsorbed with Ro red cells when tested with r'r red cells, and no reactions with the plasma adsorbed with r'r red cells when tested with Ro red cells, then the original plasma contained only anti-G.
If there is a reaction with the plasma adsorbed with Ro red cells when tested with r'r red cells, and also a reaction with the plasma adsorbed with r'r red cells when tested with Ro red cells, then the original plasma contained both anti-C and anti-D (and may also have contained an anti-G).
In all cases, however, you would give cross-match compatible C Negative, D Negative blood (you would still have to perform a serological cross-match, because there are some EXTREMELY rare donors around who are C Negative, D negative, but G POSITIVE.
As far as obstetric patients are concerned, both anti-C and anti-G usually cause far less severe haemolytic disease of the foetus and newborn (unless they have an unusually high titre), than does anti-D. However, it is important that the pregnant lady is offered prenatal and postnatal anti-D immunoglobulin prophylaxis, so that they do not get immunised against the D antigen.
I hope that this rather long and complicated post helps in some way, but, if you need to know more, please do not hesitate to ask more questions.

We do exactly the same as David.

Agreed.
:)

At UMich under Dr J Judd, a large study on anti-D reactions with gel was done in the earlier days of gel in the U.S. They concluded that anything less than or equal to 2+ by gel was suspect for weak or partial D. My poor memory thinks that the study was published in Transfusion at the time.
 
We use an Echo (2 anti-Ds) plus use a third anti-D for tube testing. If any one of the anti-Ds is less than 2+ we call the patient Rh negative and transfuse accordingly under the assumption that the patient's type is a possible weak or partial D. This is actually a pretty small number of people, so it's not sucking up the Rh negative blood supply. Someday...when molecular testing is cheaper...we will have them checked out so that we know for sure. If the patient was previously typed as Rh positive but is now called Rh negative we send a letter to the physician which explains some of the issues with Rh typing and why we are 'changing' the patient's type.

We are almost identical to Anna in the Uk.

Hi NAN47,
Any antibody that reacts at STRICTLY 37oC should be titrated during pregnancy (except, of course, for anti-D and anti-c, which, in the UK, are measured by quantification against NIBSC standards).
The titration should take place at first identification and then again at 28 weeks gestation. If the titre is less than 32, then the antibody is unlikely to cause clinically significant alloimmune haemolytic anaemia and disease of the foetus and newborn. Only rarely is an antibody titre high enough to warrant further titration after 28 weeks gestation.
The obvious exception to this is anti-K (and other Kell-related antibodies). These should be titrated when first identified, and then every four weeks to 28 weeks gestation, and then 2 weekly thereafter, until delivery. Indeed, women with anti-K who are pregnant, and who have a partner who is K+k+ or K+k- should be referred to a Specialist Foetal Medicine Unit, so that the pregnancy can be monitored by experts in the field.
It is always recommended that the previous sample is titrated in parallel with the present sample (or quantified, in the case of anti-D or anti-c), as this will show up genuine rises in the titre, as opposed to rises due to operator differences and errors and changes due to the different expression strengths of an antigen from one red cell sample to another. Where possible, the red cells used should have heterozygous expression of the antigen, although, in certain cases, this can be difficult to do (e.g., if the antibody is anti-U, it is not possible to tell whether the red cells used for the titration are U+/U+ or U+/U-).
In rare cases, titration of the previous sample, in parallel with the present sample, may show up a second specificity. For example, if the pregnant woman is known to have an anti-E, with a titre of, for example, 4 against r"r red cells, and then all of a sudden the titre goes up to 128, but ALSO shows as 128 with the previous sample run in parallel, it may be that the r"r red cells used on this occasion also express an antigen against a low prevalence antigen, against which the woman has also produced an antibody (for example, an anti-Wra).
The end point of a titration is usually given as the reciprocal of the last dilution in which agglutination can be seen with the naked eye.
We are actively re-writing the BCSH Guidelines (technical guidelines) as I post, and the complementary RCOG Guidelines (clinical guidelines) are also in their final draft.

The reason that Jk(a) variants tend to weaken the antigen expression is because of the way the amino acid residue that produces the antigen is very close to the red cell membrane and is"semi-hidden" by the third external loop (see the attached diagram).Kidd Molecule.ppt

See an earlier post here about the monolayer method ... guess that doesn't pan out the way they thought it would.  And I don't know of any 'routine' Blood Bank that runs their testing using this method.
 
In my example, since gel picks up 'everything' while the lesser sensitive method does not, we use the lesser sensitive method to perform the crossmatch.  And yes, we would transfuse antigen-negative, crossmatch compatible RBCs (crossmatch compatible with Albumin).
 
BTW: In Sunquest, we add the testing we do.  XMTS for the extended crossmatch with gel (1+, Incompatible) and XALB for the extended crossmatch with 22% Albumin (0, Compatible).  Since the patient has an Anti-E, the system requires the unit be typed/recorded as 'E-neg.'  The 'test' %TS (Transfusion Status, part of the crossmatch battery) is the determining result - the techs final interpretation.  In this case, we answer it 'OK to Transfuse' because we have determined that it is.  If we answer 'Not OK to Transfuse', the system would release the unit from the patient, i.e. no unit tag, no issuing it to the floors.
 
Of note, in the days before gel/solid phase, we probably wouldn't have seen half of these warm autos.
 
Also, a related case of interest ... for your further amusement ...
We had a 19yo female present in Sickle Cell crisis last week.  She has a positive DAT (IgG only), an Auto-Anti-E (2-3+ gel and Albumin), a 'Warm Autoantibody of Undetermined Specificity' (1+gel only) aaaand is producing Anti-e (along with a history of Anti-N).  Somewhere along the way in her short life, she got transfused with E-neg/e-pos RBCs (not all hospitals give antigen-neg RBCs proactively OR someone was thinking they should avoid the E antigen because it has a name).  This is a prime example of why we should NOT honor Auto-'identified' antibodies because of the super exposure to the 'other' antigen.  Now, this young lady will be dealing with an Anti-e the rest of her life ... through multiple transfusions (due to her Dx) and through pregnancies with an Rh-antibody.  A disservice and a shame.

Brilliant post.
The ONLY thing I would say is that monolayer methods, along with the chemiluminescence test, have proved to be less useful than was first thought. For example, anti-Inb gives a positive result, because it can cause a haemolytic transfusion reaction, but also gives a positive result vis-à-vis HDFN, which it does not cause. Some other antibodies also give "false positive" results by these methods.
That having been said, I repeat, a brilliant post!

A paper/editorial that I have quoted many times before:
Petz LD. "Least incompatible" units for transfusion in autoimmune haemolytic anemia: should we eliminate this meaningless term? A commentary for clinicians and transfusion medicine professionals. Transfusion 2003; 43 (11): 1503-1507.

It could be hyperhaemolysis, but I have my doubts.  The reason I say this is because patients with hyperhaemolysis either tend to have reactions that get worse and worse (and I include fatal reactions in this), or, for some unknown reason, tend to tolerate subsequent transfusions without turning a hair; but that does not mean that there are not patients that go down the "middle road" - it's just that they are very rare.
 
I would fall down on the side of an antibody directed against a high prevalence antigen.  However, I think I suggested a possible anti-Vel earlier in this thread.  I am now thinking more in terms of an e-variant who has made an "alloanti-e" or, worse, something like a Partial D Category 3, who has made an alloanti-D.
 
I realise that I have recommended molecular techniques a bit liberally on this site (probably because I don't have to pay for them!!!!!!! - although I do have to justify them), but I really think that full examination of the RHD and RHCE genes at a molecular level may help in this case (although it could equally be an antibody directed against another high prevalence antigen, such as anti-Jsb or anti-Fy3).  Incidentally, contrary to what many people think, homozygosity for the GATA-1 mutation and a Duffy genotype of FY*B/FY*B or FY*/FY*B does not completely exclude the possibility of forming an anlloanti-Fy3.
 
Have the clinicians tried transfusion covered by IVIG and high-dose methylprednisolone?  That may help in the short term?

A paper/editorial that I have quoted many times before:
Petz LD. "Least incompatible" units for transfusion in autoimmune haemolytic anemia: should we eliminate this meaningless term? A commentary for clinicians and transfusion medicine professionals. Transfusion 2003; 43 (11): 1503-1507.

A paper/editorial that I have quoted many times before:
Petz LD. "Least incompatible" units for transfusion in autoimmune haemolytic anemia: should we eliminate this meaningless term? A commentary for clinicians and transfusion medicine professionals. Transfusion 2003; 43 (11): 1503-1507.

I am going to be EXTREMELY controversial here, but I really don't understand why, if the cross-match is compatible, anyone would bother to type for the Cw antigen. I would be happy if someone could direct me to a RELIABLE paper that has shown anti-Cw to be clinically significant as far as an acute or delayed haemolytic transfusion reaction is concerned. I am aware of one paper in which it was claimed that anti-Cw caused hydrops in a pregnancy, but, for reasons I have given before, I do not regard this paper as reliable.
The same goes for an anti-M that is not proven to work at strictly 37oC, and many other specificities.
One only has to read the three editions of the FactsBook to see that there are numerous antibody specificities that are absolutely benign (unless your name is Lyndall Molthan - see Issitt PD. Applied Blood Group Serology. 3rd edition, 1985, Montgomery Scientific Publications, page 433. Anyone who cannot get hold of this is welcome to read my signed edition, but you'll have to give me £1 million as security, so that I get it back!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!

We, too, have dropped using the term 'least incompatible' way back when (along with 'in vivo crossmatch', I think) ... been so long, I forgot ... 1970's, 1980's?
 
1. Not all antibodies cause RBC destruction.
2. Not all antibodies that do, cause them in the same way.
3. Grade of reactivity is dependent upon the method and for some platforms, upon the tech performing the test.
4. Grade of reactivity is not proportional to the effectiveness of destruction.
 
I think the only way to determine variations of compatibility is by using monolayer methods = specialized testing.
 
Going back to 'the letter of the law': If the patient has no clinically significant allo antibodies, there is no requirement to perform an extended crossmatch. 
 
So, once this is established either by alternate testing methods (gel is very good at picking up autoantibodies) or differential absorptions, the Immediate Spin crossmatch is all that is required.  Thus, eliminating the need for conversations about 'least incompatible'.
 
If 'no clinically significant antibodies' cannot be established, it is prudent to issue antigen-negative matched RBCs to avoid the possibilities.  Some hospitals do this regardless of the current antibody status just to avoid future antibody production.
 
Of course, if there are underlying antibodies, then an extended crossmatch is required.  This should be performed with the method that was used to circumvent the auto-antibody.  Example: Assume patient has positive DAT.
Gel: All cells positive
Albumin (or LISS or PEG or whatever): Anti-E.
Use Albumin (or LISS or PEG or whatever) to perform extended crossmatch.  If this is negative and the unit is E-neg = compatible RBCs.
 
And yes, this is what we do.

Using expired antisera and panel cells is and has been standard practice for years in my experience. It has become more sound with additional QC people are now doing. If ever cited for this I would push back. Provide scientific rationale and a risk evaluation for what you are doing. These are tools used to solve problems. Without them your hands are tied and patients may not receive compatible blood. A bigger risk.
 
No matter what you call them (inspectors, investigators, assessors) they all have opinions and you may not be about to change their mind during the inspection. What they may cite you for during an inspection can be disputed in your response. The repsonse is typically reviewed by others.
 
Some of these things are just driving up the cost of health care without improving actual patient care.

A paper/editorial that I have quoted many times before:
Petz LD. "Least incompatible" units for transfusion in autoimmune haemolytic anemia: should we eliminate this meaningless term? A commentary for clinicians and transfusion medicine professionals. Transfusion 2003; 43 (11): 1503-1507.

All comments are very informative ... I'm a newbie to the site but am enjoying all the responses! It's so great to collaborate with people from other states and finding out that we are all pretty much on the same page and playing for the same team which is our patients' safety and well being.
 
I've been in this field for a little over 20 years and started out loving the job but the times have shown me the "ugly business" side of the job and frankly it is not as enjoyable as it was. You guys have made it interesting again, sounds cliché' but it's true. I'm sure I'll have more questions later but thanks again for validating what we do!

Yikes! In past years antigen typing sera could be used beyond expiration as long as there was a policy for QC. We consider them to be "rare" partially as an expense. Yes they are available from the manufacturer, but they are very expensive. Private Lear Jets are also readily available from a manufacturer, but they are rare because few people can afford them!
We will be eating some expense to get our rack in-date within the next few weeks. Our CAP window will be opening soon.
We also use expired panel cells for rule-outs.....in fact a recent CAP Survey we had to use expired panel cells to identify an anti-U....nothing in-date was helpful.
Sounds like another case of a non-technical person making rules they don't understand.

I am going to be EXTREMELY controversial here, but I really don't understand why, if the cross-match is compatible, anyone would bother to type for the Cw antigen. I would be happy if someone could direct me to a RELIABLE paper that has shown anti-Cw to be clinically significant as far as an acute or delayed haemolytic transfusion reaction is concerned. I am aware of one paper in which it was claimed that anti-Cw caused hydrops in a pregnancy, but, for reasons I have given before, I do not regard this paper as reliable.
The same goes for an anti-M that is not proven to work at strictly 37oC, and many other specificities.
One only has to read the three editions of the FactsBook to see that there are numerous antibody specificities that are absolutely benign (unless your name is Lyndall Molthan - see Issitt PD. Applied Blood Group Serology. 3rd edition, 1985, Montgomery Scientific Publications, page 433. Anyone who cannot get hold of this is welcome to read my signed edition, but you'll have to give me £1 million as security, so that I get it back!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!

A paper/editorial that I have quoted many times before:
Petz LD. "Least incompatible" units for transfusion in autoimmune haemolytic anemia: should we eliminate this meaningless term? A commentary for clinicians and transfusion medicine professionals. Transfusion 2003; 43 (11): 1503-1507.