ABID: Cold Agglutinin

[BBNBHM]

We have a issue that has been up for discussion lately at my facility regarding identifying antibodies as cold agglutinins.  

 

Per (new) supervisor, if we rule out all allos on the abid panel and our auto control is negative, we should perform a prewarm screen and if it is negative, identification is documented as "cold agglutinin".   We then perform prewarm crossmatches for the patient.  

 

I have seen this policy before at a previous facility I worked at, but this is new to some of the techs at my current facility.   And because I have been at current facility for several years now and followed the policy of the recently retired supervisor of identifying everything or sending it to the reference lab, it now makes me a little uncomfortable too.  

 

The claim by some is that there are specific tests available for identifying cold agglutinins and that we shouldn't just call them that without doing these additional tests.   Another claim is that the hematology sample would have erroneous indices if a true cold agg was present but it did not.  

 

Any input from the bb gurus on this?

 

(Ortho gel panel A and B were ran along with the initial gel screen where there were multiple 1-3+ reactions but all allos were ruled out using at least 3 homozygous cells.  Auto control negative.)

Edited February 19, 2014 by BBNBHM

[Yanxia]

I remember there had been some posts talked about this topic. If my memory not cheat me, in this kind of situation, at first we should run all the panel and cord cells at 4 degree C to see if the reaction is stronger than in room temp(22) and 37 degree. because most of pan reactive cold reactive antibodies is anti-HI or anti-I, so add the cord cells can differ it.

Not do any test just to prewarm is danger because some clinical significant antibodies like ANTI-Vel will be prewarmed out.

I don,t know if I say it right or wrong, if it is something wrong, plaese point it ou, thanks .

[tbostock]

Leger R, Garratty G. Weakening or loss of antibody reactivity after prewarm technique. Transfusion. 2003 Nov;43(11):1611-4, and the articles listed below are germane to any policy development for the use of prewarmed testing in the blood bank laboratory.

• Mallory, D: Controversies in transfusion medicine. Prewarmed tests: pro - why, when, and how - not if. Transfusion 1995 35: 268-270.
• Judd, WJ: Controversies in transfusion medicine. Prewarmed tests: con. Transfusion 1995 35: 271-275.

 

I'm not a fan of prewarming.  I think there is a danger of prewarming away alloantibodies.  You should also prove that there is cold reactivity first, and use other methods for detecting alloantibodies.  We only use prewarming here as a very last resort.

[SMILLER]

Years ago, we used to "verify" colds by doing the 4 C incubation thing, but we decided it is unecessary. A "cold" here is either significant (if it reacts at 37 or coombs) or insignificant. So we report them as such.

Scott

[R1R2]

I understand your concern and would first want to identify a cold by running appropriate tests before jumping right to prewarm and assuming that because reactions went away that it was due to a cold.  Many years ago when I was a new supervisor, I observed exactly what you described and was bothered by it.   I reviewed past workups that were identified as cold abs and found that many were really wishy washy anti M's.   I did find one Fya  that was reacting sporadically with only double dose cells.   The way they performed antibody id was quickly changed to stop the practice of prewarming away unexplained reactions. 

Edited February 19, 2014 by R1R2

[carolyn swickard]

I agree with R1R2 - at least show that there is a cold there (at least do a cold screen) before assuming an unclear panel is just a "cold".  Always check the heterozygous vs. the homozygous patterns first too.

 

I have seen way too many weak antibodies over the years that did not fit a perfect panel pattern, but were anything but "just a cold".  Prewarming is a weakened reaction system without any help from the current enhancement medias and it is perfectly capable of missing a bunch of stuff.  I did have a reference lab tell me once that you could "prewarm" with enhancement medias too - "just prewarm the media too" and we have done some of that over the years, but not everyone does. 

[Justina]

Just like everyone else I too do not like the idea of calling unclear reactivity a cold based on a prewarmed screen/panel. Not every antibody reacts text book and I feel more comfortable giving non-prewarmed cxm compatible units than prewarmed cxm compatible. I have also found that after repeating other techs work...they sometimes miss a weak positive reactions. So better play it on the safe side.

[Townsend]

We also avoid any pre-warming if at all possible.  If there are non-specific reactions that don't seem to follow any particular pattern (in gel, our routine method), we will do a "cold screen" (just incubation of the screen cells at 4 deg C).  If positive at 4 deg, we call it a cold agglutinin or cold auto if AC is positive.  We still rule out all common antibodies using gel.  No further ID of any cold agglutinins (i.e. anti-I/IH) unless it is requested due to clinical signs and symptoms of hemolysis.  At that point, further cold ID testing would be sent out.

[Malcolm Needs ☆]

Townsend, I just cannot see what difference the cold agglutinin specificity makes due to the clinical signs and symptoms of haemolysis (with the POSSIBLE exception of anti-Pr - and I'm not convinced about that either, and, of course, anti-P in the case of PCH, but then the 2-stage DL test is better for that).

Of paramount importance is the thermal amplitude of the antibody.

[SMILLER]

I am still not sure what is the use of "verifying" a cold by doing a cold panel everytime one is suspected.  We don't even screen units when the patient has non-37/AHG anti-M anymore.  If there are no significant antibody-antigen problems at 37 or coombs, what difference does it make?  When are you going to transfuse a patient at 4 C?

 

Is it something to do with documentation or using a blood warmer?

 

Scott

Edited February 23, 2014 by SMILLER

[Malcolm Needs ☆]

Could NOT agree more SMILLER.

There is being thorough, and there is being WAY over the top.

[Brenda K Hutson]

I have taught through the years that one should "never" assume something is a Cold Agglutinin unless they prove it (with a cold panel; i.e. Screening Cells, Auto, Cord and whatever Reverse Cells the patient is Negative for). I even have a story of a place with a similar protocol to what you discuss (though much worse).

When I was a Reference Lab Supervisor, we received a "very" hemolyzed specimen on a patient who had a hemolytic transfusion reaction. The Hospital sent their panel work. They did run a panel; and it was a perfect pattern of an Anti-E; but their Policy (and I can only hope they misunderstood it) was that they should try to perwarm it away and if it went away, it was not significant. A week later, we received another hemolyzed specimen from the same Hospital (different patient). This time the patient had a perfect E and c; but again, they managed to prewarm them away (which is another pet peeve of mine.....trust me, I have seen many clinically significant antibodies prewarmed away...by good Techs.; even in large, prestigious Medical Centers....so I tend to be very anti-prewarm except in the right hands). Anyway, I called their Medical Director and told her they needed to "cease and desist" with prewarming before they killed someone (not to mention, teach their Techs. how to perform basic Antibody ID).

Your Policy is not "that" bad in that your supervisor is saying you have first ruled-out major alloantibodies. But let me tell you another problem (sorry to belabor the point). If you use GEL, know that I have seen "many" instances of Kidd group antibodies that not only do not react with any heterozygous cells; but do not even react with "all" homozygous cells. So just saying you ruled everything out, doesn't necessarily mean you did. Not only have I seen that in Hospitals I have worked in; but also in work-ups sent to us at the Reference Lab from Hospitals that used GEL but did not know of this potential problem.

So, I have also taught through the years that even when you think you have ruled everything out, you need to look at your positive reactions and see "what do all of those cells have in common?" You might run into some surprises.

Brenda

[Malcolm Needs ☆]

Don't necessarily agree with all of your post, but, that notwithstanding, an excellent post Brenda, which should be read by all and acted upon by all.

[Brenda K Hutson]

 Would like to hear what part you don't agree with.....I am always open to discussion.

Thanks Malcolm,

Brenda

Don't necessarily agree with all of your post, but, that notwithstanding, an excellent post Brenda, which should be read by all and acted upon by all.

 

[Dansket]

We have a issue that has been up for discussion lately at my facility regarding identifying antibodies as cold agglutinins.  

 

Per (new) supervisor, if we rule out all allos on the abid panel and our auto control is negative, we should perform a prewarm screen and if it is negative, identification is documented as "cold agglutinin".   We then perform prewarm crossmatches for the patient.  

There was a poster at the most recent Annual Meeting of the AABB suggesting that running an enzyme panel may be useful in your situation (all allo's ruled out in gel panels with one or more reactive panel cells).

 

I discarded prewarmed testing a long time ago, for the same reasons others have posted.  Based on your scenario, I would issue -IgG crossmatch-compatible blood. 

 

What does your supervisor recommend when the prewarmed antibody screen is positive?

Edited February 24, 2014 by Dansket

[Malcolm Needs ☆]

Very, very little Brenda (and I am probably being overly pedantic).

It is just that antigens cannot be either homozygous or heterozygous; only genes can be. Antigens can show homozygous expression or heterozygous expression, but they cannot be homozygous or heterozygous.

I agree with 99.9% of what you say.

[Brenda K Hutson]

Ok, while you look up EGA...I will look up pedantic! LOL

I stand corrected......genes.

Thanks for responding.

Brenda

[Brenda K Hutson]

 

Ah, but looking back at my post; I wrote "cells" not "Antigens;" so seems like more of a reflection of just interpeting the antigram in reviewing one's work (or me just being pedantic).

Brenda

Very, very little Brenda (and I am probably being overly pedantic).

It is just that antigens cannot be either homozygous or heterozygous; only genes can be. Antigens can show homozygous expression or heterozygous expression, but they cannot be homozygous or heterozygous.

I agree with 99.9% of what you say.

 

[Malcolm Needs ☆]

Ah, but red cells do not have a nucleus, and so thay cannot have genes!!!!!!!!!!!!!!!

[Joanne P. Scannell]

I agree with the cautions ... here is what we do for your consideration:

 

Our pretransfusion testing includes a test we call 'O CELLS'.  (This is a pool of the 3 cells from the 3% Antibody Screen).  It essentially replaces the old I.S. phase from tube testing so it tells us if there are 'cold agglutinins' or 'rouleaux' that would need to be considered during crossmatching.

Test: 2 drops patient plasma + 1 drop 'O cells' ... mix, spin, read, grade and record. (Immediate Spin Phase). 

If results are questionable, incubate at 22C for 15 minutes and respin (22C Inc Phase).

 

The result is helpful information when we see mixed reactivity in the Antibody Screen (using 37C with gel) ... both cold agglutinins and rouleaux can look the same in gel.

 

Because gel is somewhat acidic, we have seen many more Anti-M's than we saw using tube testing.  If we identify Anti-M and the Ocells were also positive due to cold agglutinin, we rerun plasma vs M-pos cells prewarmed to 37C to determine the thermal amplitude of the Anti-M. 

 

No reactivity at 37C = Cold Agglutinin: Anti-M. (No antigen testing but use a blood warmer.)

Reactivity at 37C = Anti-M (Reactive at 37C) ... this one is classified as clinically signficant.

 

Ditto for others, e.g. Lewis.

 

It's simple and it works for us.

 

 

[Joanne P. Scannell]

I have taught through the years that one should "never" assume something is a Cold Agglutinin unless they prove it (with a cold panel; i.e. Screening Cells, Auto, Cord and whatever Reverse Cells the patient is Negative for). I even have a story of a place with a similar protocol to what you discuss (though much worse).

When I was a Reference Lab Supervisor, we received a "very" hemolyzed specimen on a patient who had a hemolytic transfusion reaction. The Hospital sent their panel work. They did run a panel; and it was a perfect pattern of an Anti-E; but their Policy (and I can only hope they misunderstood it) was that they should try to perwarm it away and if it went away, it was not significant. A week later, we received another hemolyzed specimen from the same Hospital (different patient). This time the patient had a perfect E and c; but again, they managed to prewarm them away (which is another pet peeve of mine.....trust me, I have seen many clinically significant antibodies prewarmed away...by good Techs.; even in large, prestigious Medical Centers....so I tend to be very anti-prewarm except in the right hands). Anyway, I called their Medical Director and told her they needed to "cease and desist" with prewarming before they killed someone (not to mention, teach their Techs. how to perform basic Antibody ID).

Your Policy is not "that" bad in that your supervisor is saying you have first ruled-out major alloantibodies. But let me tell you another problem (sorry to belabor the point). If you use GEL, know that I have seen "many" instances of Kidd group antibodies that not only do not react with any heterozygous cells; but do not even react with "all" homozygous cells. So just saying you ruled everything out, doesn't necessarily mean you did. Not only have I seen that in Hospitals I have worked in; but also in work-ups sent to us at the Reference Lab from Hospitals that used GEL but did not know of this potential problem.

So, I have also taught through the years that even when you think you have ruled everything out, you need to look at your positive reactions and see "what do all of those cells have in common?" You might run into some surprises.

Brenda

 

I think pre-warm testing got it's bad reputation because in the 'old days', when we were running antibody screens that included the IS/22C phases and used polyspecific AHG, if the test was positive at AHG and also at room temp, we would rerun the AHG phase using 'Prewarm Technique' to see if we were seeing a complement fixation due to cold agglutinins or a real IgG antibody. 

 

In those days, Prewarm Technique meant using NO enhancement; albumin wasn't allowed because it was said to enhance complement uptake, which is what we were trying to avoid.  The change from Albumin to 'nothing' lowered IgG uptake, hence the sensitivity ... so yes, of course we were risking losing sight of clinically signficant warm antibodies.

 

The world has changed since then ... we don't use polyspecific AHG anymore, most places don't test at room temperature anymore and prewarming is merely just that - same test only at 37C before you mix plasma with cells.

 

In fact, if using automation (e.g. ProVue), the antibody screens are all pretty much prewarmed because they are sitting in the 37C incubator for at least a few minutes prior to adding plasma/reagents.  So, we are testing 'prewarm' by the thousands daily ... and nobody is fearful about missing antibodies due to prewarming.

 

Prewarming is no longer 'lowering sensitivity' if that is the only parameter that is changed.

 

I agree ... this hospital you cited was mistaken and hopefully has corrected their workup plan:  'Prewarming' away an antibody that is clinically signficant is a strange policy ... similar to 'just shake it harder' or 'repeat until negative'.  HOW were they 'prewarming'?  Were they using the old 'Prewarm Technique'?

 

And yes, thank you for stressing ... we, too, have seen plenty of antibodies that react with ONLY homozygous cells, even in gel. (So please no talk out there about 'eliminating with 3 heterozygous').  

 

And I agree with 'look at what positives have in common' ... we found that PEG will demonstrate a nicer Anti-Jka than gel sometimes, so if you get a hint of Kidd ... or a hint of anything significant, try PEG. 

 

More evidence that we have to carry a 'toolbox', not depend on one reagent/platform/technique. 

 

And that we cannot take all the old 'rules' and apply them to all the new processes.

Edited February 25, 2014 by JPCroke

[Joanne P. Scannell]

Townsend, I just cannot see what difference the cold agglutinin specificity makes due to the clinical signs and symptoms of haemolysis (with the POSSIBLE exception of anti-Pr - and I'm not convinced about that either, and, of course, anti-P in the case of PCH, but then the 2-stage DL test is better for that).

Of paramount importance is the thermal amplitude of the antibody.

Love it!

[Brenda K Hutson]

 

I think pre-warm testing got it's bad reputation because in the 'old days', when we were running antibody screens that included the IS/22C phases and used polyspecific AHG, if the test was positive at AHG and also at room temp, we would rerun the AHG phase using 'Prewarm Technique' to see if we were seeing a complement fixation due to cold agglutinins or a real IgG antibody. 

 

In those days, Prewarm Technique meant using NO enhancement; albumin wasn't allowed because it was said to enhance complement uptake, which is what we were trying to avoid.  The change from Albumin to 'nothing' lowered IgG uptake, hence the sensitivity ... so yes, of course we were risking losing sight of clinically signficant warm antibodies.

 

The world has changed since then ... we don't use polyspecific AHG anymore, most places don't test at room temperature anymore and prewarming is merely just that - same test only at 37C before you mix plasma with cells.

 

In fact, if using automation (e.g. ProVue), the antibody screens are all pretty much prewarmed because they are sitting in the 37C incubator for at least a few minutes prior to adding plasma/reagents.  So, we are testing 'prewarm' by the thousands daily ... and nobody is fearful about missing antibodies due to prewarming.

 

Prewarming is no longer 'lowering sensitivity' if that is the only parameter that is changed.

 

I agree ... this hospital you cited was mistaken and hopefully has corrected their workup plan:  'Prewarming' away an antibody that is clinically signficant is a strange policy ... similar to 'just shake it harder' or 'repeat until negative'.  HOW were they 'prewarming'?  Were they using the old 'Prewarm Technique'?

 

And yes, thank you for stressing ... we, too, have seen plenty of antibodies that react with ONLY homozygous cells, even in gel. (So please no talk out there about 'eliminating with 3 heterozygous').  

 

And I agree with 'look at what positives have in common' ... we found that PEG will demonstrate a nicer Anti-Jka than gel sometimes, so if you get a hint of Kidd ... or a hint of anything significant, try PEG. 

 

More evidence that we have to carry a 'toolbox', not depend on one reagent/platform/technique. 

 

And that we cannot take all the old 'rules' and apply them to all the new processes.

I am referring to prewarm testing only at 37C and AHG (separate sets of tubes set up). There are still a lot of details that can be done incorrectly if not experienced (prewarming of plasma/serum; temp. of saline which is the big one; method of washing, decanting, then adding IgG AHG). I still think it deserves a bad rap except in the right hands (but that is just my bias). I much prefer RESt.

I don't know that I would "think of" GEL ProVue Testing as Prewarming….we certainly still see plenty of cold agglutinins in GEL (I think just based on the methodology of the GEL column). Don’t know…

Brenda

[Brenda K Hutson]

 

Ok, so "now" you are being pedantic! LOL (would put a smiley face but still haven't figured out how to use this website; i.e. for replies). Ugh

Brenda

 

Ah, but red cells do not have a nucleus, and so thay cannot have genes!!!!!!!!!!!!!!!

 

[Mabel Adams]

I see several people are channeling for John Judd here.  He was asked to fill in at the last minute for another speaker at the last AABB meeting and he managed to get in a reminder to not do the pre-warmed technique (which most of us define as a saline only tube method where reagent cells and plasma sample are warmed before combining and washed with 37 degree saline before addition of anti-IgG).  He used to refer to it as the pre-fried technique because people had a tendency to use ever warmer saline if they couldn't get the antibody to "go away" by using 37 degree saline.  Most of the missed antibodies in the study he published were due to the less sensitive technique of saline testing.  The thing is, you very seldom need to do it.  If you can rule out all of the usual suspects and get AHG crossmatch compatible units by a more sensitive method that is what you should do.  Save pre-warmed for patients with a really potent cold where you have no other choice and then don't "fry" the cells.  Then you don't often need to fuss with cold panels and the like because their cold antibody will probably jump out of the tube at you--or at least interfere with the reverse type.

 

Malcolm, note that I said "often".  (and I put the period after the quote marks like you Brits do.    )