saline replacement in serum discrepancy

[larevalo]

How do you perform saline replacement when there is serum discrepancy due to rouleaux formantion?From the last step,2 drops of saline is added and resuspend it with red cells does it need to read the agglutination macroscopically?(because iso-antibodies are IgM antibodies) AABB Technical manual did not estates if microscopic or macroscopic.Any help?

[Yanxia]

I read it macroscopically, and for microscopical I will used in not serious rouleaux because it looks like coin upon coin.

[larevalo]

I read it macroscopically, and for microscopical I will used in not serious rouleaux because it looks like coin upon coin.

Hi shilly,thanks for your reply,only i want to clarify if you see the reaction macroscopically and the results is negative,but your microscopic reading rouleaux formation is not completely ruled-out, are you going to report this as negative?

[khalidm3]

If Rouleaux persist try saline replacement. Mostly it is successful for micro or macro readings. Centrifuge reaction tube, remove serum/Plasma without disturbing the red cell button and add equal volume (2 drops) of saline, Mix and read. Preferably use negative control (O cells) for reverse.

Edited November 13, 2010 by khalidm3

[L106]

Larevelo, I think that's up to your Lab to decide what your routine procedure will be. Off the top of my head, I can't think of any test that AABB dictates must be read microscopically (except FMH Screening.) So I think that most Labs that read their tests only macroscopically (most likely with a view mirror) will require that tests will be read the same way after saline replacement.

On the other hand, if my generalist staff (that rotate all departments and may not be in Blood Bank for several weeks) want to peek at the test under the microscope after the saline replacement to be sure they aren't missing a weak reaction, I can live with that.

Just my thoughts on the subject.........Donna

[COTTONBALL]

I agree with L106. As you know, you have to scope it to confirm the "stacks of coins" and so, to take a peak at it (microscopically) after saline replacement, should not do any harm. Some weak reacting, cold antibodies, plus Anti-A1 (reverse typing)may be lurking. I would follow my written procedures.

[Eagle Eye]

we read under microscope to see if rouleaux is present and after saline replacement we read under microscope to confirme negative result.

[larevalo]

If Rouleaux persist try saline replacement. Mostly it is successful for micro or macro readings. Centrifuge reaction tube, remove serum/Plasma without disturbing the red cell button and add equal volume (2 drops) of saline, Mix and read. Preferably use negative control (O cells) for reverse.

Thanks khalidm3.

[larevalo]

Thanks Donna,I think you are right .I'll let my Specialist decides which routine procedure we are going to apply.

[khalidm3]

Since we shifted to Gel the problem of rouleau is minimized. Microscopy is not recommended for most of our procedures, but it helps to rule out doubts. I am in favour of Gel or automated micro-plate, the procedures are hygienic and easy to read, uniformed.

[Yanxia]

Hi shilly,thanks for your reply,only i want to clarify if you see the reaction macroscopically and the results is negative,but your microscopic reading rouleaux formation is not completely ruled-out, are you going to report this as negative?

Hi, larevalo.I am sorry for reply so late, because my computer has problem those days, I just can read the post but can't reply it.

Does you mean the salin replament is also coagulation and microscopic review is rouleux , in this condition I will report is as rouleux, because I have seen some very strong so saline replacement do not show neg .

[Yanxia]

Since we shifted to Gel the problem of rouleau is minimized. Microscopy is not recommended for most of our procedures, but it helps to rule out doubts. I am in favour of Gel or automated micro-plate, the procedures are hygienic and easy to read, uniformed.

Khalidm3, In my routine work, I think gel is not suitable to used in rouleux specimen, because the protein abnormal will give the screening cells all positive.

This is just my gel card, I want to know if you have the different gel card.

[larevalo]

Hi, larevalo.I am sorry for reply so late, because my computer has problem those days, I just can read the post but can't reply it.

Does you mean the salin replament is also coagulation and microscopic review is rouleux , in this condition I will report is as rouleux, because I have seen some very strong so saline replacement do not show neg .

hi Shily, got your response,thanks.

[khalidm3]

Khalidm3, In my routine work, I think gel is not suitable to used in rouleux specimen, because the protein abnormal will give the screening cells all positive.

This is just my gel card, I want to know if you have the different gel card.

I do not remember any unsolved case of rouleau in reverse, since we shifted to Gel, from DiaMed. We have an average of 27000 blood donor's, 30000 type and screen/cross match, and 30000 specimens for blood grouping per year. Some time we faced problem with cold autoimmune/antibody identifications and if we were unable to solve, refered such cases to some other centres.

[larevalo]

Hi khalidm3,thanks for the info.,we do have reference lab.but before we send to them we have to find our way to rule it out .For ABO/Rh typing we are using tube method only.Both tube & gel methods are applied mainly for ABS & ABI .

[khalidm3]

Hi khalidm3,thanks for the info.,we do have reference lab.but before we send to them we have to find our way to rule it out .For ABO/Rh typing we are using tube method only.Both tube & gel methods are applied mainly for ABS & ABI .

Larevalvo, sure we have to work out and use all available sources to resolve a problem before refering. We are now 100% on Gel and soon we are shifting our one area, mostly donors to Tango for which we are hanged with the BioTest distributor service here.

[Brenda K Hutson]

If the rouleaux is such that the initial testing looks "rough" macroscopically upon resuspension, I then look at it microscopically to be confirm that it is rouleaux. So after performing the saline replacement, I again look both macroscopically and microscopically because I want to ensure the saline replacement really did get rid of the problem (i.e. make sure it was not a mixture of rouleaux and agglutination).

Brenda Hutson, CLS(ASCP)SBB

How do you perform saline replacement when there is serum discrepancy due to rouleaux formantion?From the last step,2 drops of saline is added and resuspend it with red cells does it need to read the agglutination macroscopically?(because iso-antibodies are IgM antibodies) AABB Technical manual did not estates if microscopic or macroscopic.Any help?

[khalidm3]

If the rouleaux is such that the initial testing looks "rough" macroscopically upon resuspension, I then look at it microscopically to be confirm that it is rouleaux. So after performing the saline replacement, I again look both macroscopically and microscopically because I want to ensure the saline replacement really did get rid of the problem (i.e. make sure it was not a mixture of rouleaux and agglutination).

Brenda Hutson, CLS(ASCP)SBB

I do the same too

[Brenda K Hutson]

Aahh...please forgive my poor grammar (and/or spelling) once again....i.e. "to be confirm." I have got to take time to review before posting!

Brenda

I do the same too

[khalidm3]

Aahh...please forgive my poor grammar (and/or spelling) once again....i.e. "to be confirm." I have got to take time to review before posting!

Brenda

Grammar and spelling do not matter much, Your concept is very clear and you expressed it excellently.

[larevalo]

If the rouleaux is such that the initial testing looks "rough" macroscopically upon resuspension, I then look at it microscopically to be confirm that it is rouleaux. So after performing the saline replacement, I again look both macroscopically and microscopically because I want to ensure the saline replacement really did get rid of the problem (i.e. make sure it was not a mixture of rouleaux and agglutination).

Brenda Hutson, CLS(ASCP)SBB

Hi Brenda, got it clear.Thanks.

Which do you apply first in serum typing discrepancy,(under microscope read as true agglutination in combination of rouleaux) Saline replacement or anti-A1 Lectin for the presence of unexpected anti-A1 in reverse? or both?

[Brenda K Hutson]

I do the Saline Replacement first for a couple of reasons:

1. "Stacks of coins" are not always seen just as long strands; they can curl up on themselves and lead the less experienced

person to think it is agglutination.

2. An A subgroup with Anti-A1 is more often a stronger reaction with A1 cells than what one sees with rouleaux

3. An A subgroup with Anti-A1 is not seen as frequently as rouleaux (at least not in my experience; but that also depends

somewhat on your patient population)

4. There are other considerations in addition to rouleaux or anti-A1 with unexpected reactivity with A1 cells; i.e. Cold

Agglutinin; or an Antibody which reacts at cooler temps. such as M for example. But if one of these is suspected, I am

NOT one to say "try prewarming and if it goes away, it was a cold." Bad idea (but unfortunately, a more common

approach than I even like to think about)! You need to first prove you have a cold (which then may or may not have a

specificity).

Just my thoughts...

Brenda Hutson, CLS(ASCP)SBB

Hi Brenda, got it clear.Thanks.

Which do you apply first in serum typing discrepancy,(under microscope read as true agglutination in combination of rouleaux) Saline replacement or anti-A1 Lectin for the presence of unexpected anti-A1 in reverse? or both?

[khalidm3]

Brenda I suggest to use Negative Control (O Cells) with reverse especially when there is a discrepancy to rule out unexpected antibodies. We should not shift to cold until it is proved

Edited November 19, 2010 by khalidm3

[larevalo]

Hi Brenda, got it clear.Thanks.

Which do you apply first in serum typing discrepancy,(under microscope read as true agglutination in combination of rouleaux) Saline replacement or anti-A1 Lectin for the presence of unexpected anti-A1 in reverse? or both?

Hi Brenda,what a big help!thats the advantage of being a member of this forum,I do appreciate to all the people who had shared and answered to my thread,thanks blood bank talk:)...P.S. anti-M is eliminated, because antibody screening is clear negative both in tube and gel method.

[larevalo]

yes ,khalidm3 i agree that we need to have negative control in reverse by using o cells.