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Posts posted by exlimey
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22 hours ago, saralm88 said:
Hi friends!
I just wanted to get a feel for what different labs are using. We have the Echo and Neo (new) and have been using LISS for screens and xm. I am switching to using PEG because Immucor says it is the best to use when having the machines. What does everyone else use out there?
I hope everyone is having a great summer! I look forwards to hearing from you - hopefully I can get on here more and give more input of my own!
Thanks!
PEG-IAT is arguably the most sensitive tube test currently in widespread use. For this reason you're more likely to see concurrence with your Echo/Neo results if you use PEG for supplementary testing, i.e., the sensitivity of the two assays are perhaps the closest (LISS being less-sensitive). However, there's still a chance that the Echo/Neo will detect something that is not detected in PEG (or LISS).
I wonder what Immucor would say if you decided to use another manufacturer's PEG reagent ?
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On 7/6/2018 at 4:35 PM, Cliff said:
Folks, please read this carefully, I suggest he is saying it is acceptable to call him Malcolm, as compared to Mr. Needs - hence informal. He did not suggest he would find it acceptable to refer to him as a tosser, wanker, or some other (really cool sounding British) pejorative.
I need to move to England, if only for a short time. American English is so boring.
I think the term "Chav" has recently become popular. Even more recently, calling someone "a right old Neymar" is not very flattering.
As the Irish writer George Bernard Shaw once said: "England and America are two countries divided by a common language."
- Malcolm Needs and AMcCord
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15 hours ago, TreeMoss said:
The physicians would use cold cardioplegic solution -- 4 degrees - when putting the patient on bypass.
Wow. Thank you for that information. That certainly could influence the concern some of the medics demonstrate. Is the surgical room also chilled ?
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10 hours ago, rravkin@aol.com said:
Hi Scott and Exlimey, perhaps the idea of a hemolytic reaction is remote when considering cold agglutinins but if the patients body temp and OR temp are below normal body temp and room temp respectively, and, if the cold agglutinin is strong enough, either by way of concentration therein of bonding strength, does the cold agglutinin not have the capability of causing rbc agglutinins to form and remain stable long enough to cause damage to the microvasculature in the same manner as with Lupis Erythmatosis.?
That is exactly the theoretical risk that concerns the medical staff, but in my non-medical, laboratory-based opinion, the risk is extremely low. Extreme testing protocols (below 30 C) for cold-agglutinins are rarely informative, often having very specious clinical relevance. Does anyone really know what the results mean ? How high must a titration be to be significant ? If you look hard enough, you can find cold-reactive autoantibodies in most people, hence why routine testing protocols now deliberately avoid test phases below 37 C. Modern, super-sensitive test systems (PEG-IAT, CAT) don't even allow tests below 37 C and openly admit that IgM antibodies may not be detected (typically the form that "colds" take). Even with these "deficiencies" they still are licensed/approved for antibody detection and ID.
If a patient is in such a dire situation that they're undergoing radical surgery, with the selective use of hypothermia and/or by-pass procedures, the least of their worries is a cold agglutinin.
The easy fix to the transfusion of "cold blood" is a blood warmer, but obviously this would be contraindicated during hypothermic processes.
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Perhaps I'm a little naive, but I find some of the "old time" logic somewhat illogical. I appreciate that a unit of red cells being transfused would potentially be "cold" - 1 - 6 C at the start of infusion, i.e., might cause a cold-agglutinin issue, but almost immediately, the infused portion would equilibrate to the temperature of the circulating blood. Additionally, the unit itself would start to warm-up to room temperature.
Certainly additional problems could arise from "by-pass" procedures, but are the devices\pumps "cold" - 1 - 6 C ?? I suspect they operate at room temperature, nowhere close to refrigerator temperatures.
After all that rambling, I meant to say that I don't why anyone would test "cold autoantibodies" at temperatures below that of typical (surgical) rooms. However, I'm sure there is a a whole library of circumstantial, anecdotal evidence supporting such extreme testing protocols.
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On 6/22/2018 at 3:22 PM, richj said:
Hello
What are the pros and cons of using IgG vs AHG when performing a tube crossmatch or tube screen assuming that the tube method is a back up method to MTS gel.
Is it possible to miss a complement binding IgM antibody early on by using IgG only. The standards seem to indicate that IgG only is required.
Thanks.
Richard
Does the MTS gel card you typically use contain polyspecific antiglobulin reagent (anti-IgG + anti-complement) or does it just contain anti-IgG ? I think most users are using anti-IgG cards, and if that is the case, they're already dealing with the "Is it possible to miss a complement binding IgM antibody early on by using IgG only." issue.
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20 hours ago, SBBSue said:
I've been told that the concentration of DTT in RESt is different that what is used for treatment of reagent cells. But that sure would have been convenient.
RESt = Rabbit Erythrocyte Stroma - basically stabilized red cell membranes from rabbits. There is absolute no DTT in RESt.
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I just answered this question.
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My ScorePASS
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I suspect that routine use of enzyme-treated cells (in IAT) by "Non-reference Laboratory Staff" would cause more confusion than it would solve. Even the largest, most proficient hospital laboratory doesn't have high caliber serologists available on all shifts. I would suggest that tests with enzyme-treated cells be restricted to more difficult serological pictures, e.g., post-transfusion hemolysis without obvious cause (read "anti-Jka or anti-Jkb"), or for investigation of antibodies to high-incidence antigens.
I also suspect that many of the "enzyme-only" specificities have a major IgM component - notoriously difficult to detect by CAT (gel).
Just my two cents/pennies.☺
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4 hours ago, Ensis01 said:
My understanding is that the DAT negative saline control is only required when the patient has a positive poly, IgG and compliment DAT. The DAT saline control, when negative, rules out pollyagglutination (either microbial or acquired). Therefore if the patient's Poly or anti-IgG or anti-C3b,-C3d DAT is negative; pollyagglutination is ruled out and the saline control is not needed.
Polyagglutination (one L) is a phenomenon demonstrated by some red cells in the presence of most normal human sera. It has little to do with a positive DAT. Perhaps you mean "spontaneous" or "non-specific" agglutination ?
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19 hours ago, cswickard said:
Yes, but it is also - according to years of CAP survey results - routinely 1-2 endpoints higher on every titration run. So, make sure you are telling your physicians that your facility is doing titers with gel and that the actionable titers for their patients might be at higher endpoints than those historically with tube testing.
I wonder if the inspector would have been OK with procedures with this kind of control if it was used when there was no retained sample (no control avail) and not used when the new specimen was run in parallel with it's retained pair (internal control avail)?? I guess we will have to see, because that is what we are going to try doing.
You bring up some interesting points and I agree with your position.
Certainly "the literature" regarding the clinical importance of titration results is confusing. Most of the original work was done on anti-D, using an unorthodox test protocol (I believe the titrant was a high concentration of BSA), but there did seem to be some correlation between titration strength and clinical impact. Workers attempted to shoe-horn other specificities into the same program, with mixed success. Now, today, as you point out, the gel test is becoming a routine way to measure antibody strength. I don't think anyone honestly knows what the titration end-points mean, since the modern results are difficult to interpret/compare to the older literature. Time will tell.
I think people have been caught in a little trap with regard to the controls needed for titrations. It is very unusual to have two sequential samples in a clinical assay, even rarer that one of said samples has been stored (frozen). Consider a simple CBC.....many patients with extended hospital stays have multiple tests performed. Their last samples are not run in parallel with the current sample, yet the results are considered valid because the instrument's controls performed as expected - this is the equivalent of using material with a known potency as a control for patient sample testing. The art is in selecting your control.
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21 hours ago, SMILLER said:
Well, it may seem subjective to YOU...
The particular problem we had was with a few panel cells on a new panel coming up only slightly hazy--not even a weak 1+. (These are R1R1 cells that should be clearly positive with our usual QC anti-sera which contains anti-D and ant-e. So that is what is under investigation.) Ortho said that we should be seeing a definite 1+ at the least. We are used to seeing 2+ or 3+ with these gel panel cells.
Scott
Your results suggest to me that your QC reagent is deteriorating, maybe, perhaps, subjectively speaking, of course.☺
I am well aware that manufacturers test every cell for every antigen and that the results must meet a certain threshold. However, as Malcolm suggests, "decent strength" can be a very elusive target. One direct test with undiluted antisera (or a strategically-diluted sample) does not always give an honest indication of antigen strength.
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38 minutes ago, SMILLER said:
Yeah, I wondered about that hetero/homozygous D thing as well... I will also mention that according to Ortho, Panel cells are all tested with specific antisera for each antigen for appropriate reaction strength before being released for shipping.
Scott
Yeah, that's one of the many fuzzy statements found in package inserts. I suspect that it means they perform a test to detect antigens and the reaction has to meet a certain grade. I doubt that they test the strength of antigens by titration or by another other more exotic means (FACS).
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16 minutes ago, Malcolm Needs said:
(as there is no such gene as RHd, there cannot be a heterozygous form - exlimey!) - I stand corrected ! Thank you.
Lastly, one would hope that, if the panel you are using is a commercial panel, they would have done something (perhaps a fluorescence-labelled anti-D and a FACS) to ensure that no red cells are chosen with a particularly low number D antigens expressed. - Not a chance of that happening.☺
The exons involved in the RHD gene leading to Partial D Category IV, are 2, 3 and 7 (exlimey), but even these express around 9, 000 D antigens. I was on the right track.
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9 minutes ago, gagpinks said:
We did 4 A Rh D negative and they all were compatible. Waiting for eluate result. Patient had 1 B neg platelet.
That conflicts with you original post: "Set up 4 units of A Pos unit and found to be incompatible. Then we set up 3 A Rh D negative units and found to be incompatible."
I'm confused.
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Two thoughts:
1. The other D+ cells on the panel probably have "homozygous expression", whereas the Ro donor may be a single dose. You may just be seeing a dosage effect.
2. The Ro cell may be DcatIV, which is missing a few epitopes found on normal expressions of D antigen (I don't remember the details). If the donor is heterozygous (single dose), expressing only the DcatIV gene, the anti-D in the patient may not have the ability to react with the modified expression.
If Malcolm Needs reads this, I'm sure he'll correct my terminology deficits to the more modern versions.
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On 3/21/2018 at 7:08 AM, gagpinks said:
Hi
Patient blood group is A Rh D positive and antibody screen negative. Set up 4 units of A Pos unit and found to be incompatible. Then we set up 3 A Rh D negative units and found to be incompatible. Therefore panel was set up and found to negative in all cell line. We set-up 4 unit O Rh D negative and found to be COMPATIBLE. However DAT is positive in IgG 2+. But why was cross match was incompatible in A red cell but COMPATIBLE with O red cell.
On another track......why the switch to Rh-negative units ?
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1 hour ago, cheru26 said:
Have you looked into the possibility of patient having low incident antibody? Low incident antibody are usually picked up during crossmatch.
This doesn't fit the pattern for an antibody to a low incidence antigen - in this case, all 7 group A units are incompatible and the 4 group O units are compatible.
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Here's my 2-cents, 2-pennies, or 2-any other small denomination coins:
First, I am NOT a regulatory expert, but I am familiar with assay development and validation.
All assays should be controlled in some fashion, to give the practitioners some confidence that the results are valid AND that batch-to-batch variation is limited. In a titration, especially a serological titration, this is a little more difficult than having one or two pass/fail samples that are typically included in many laboratory tests.
If a control is needed for a titration, it doesn't need to be the same specificity as the test antibody (as Malcolm highlights). Ideally, yes, but not necessarily. It does need to be reliable/robust and give the same end point each time, even with the acknowledged variations in serological tests (reagents, test cells, techs, etc.). Tube testing is notoriously variable, while gel testing is believed to reduce some of those nuances. As Malcolm suggests, it might be necessary to have a control for an IgM titration (ABO) and/or a different one for an IgG titration. At the very least, the end-points may be different. An IgM control might be as simple your routine anti-A reagent; a simple IgG control might be an IAT-reactive anti-D or other specificity. A clever option might be a control that contains both - an IgM component and an IgG component.
If a test system is adequately controlled each time (and passes), in my opinion, there is no reason to routinely perform parallel testing of successive patient samples. Retention samples might be useful for investigatory reasons if/when a patient's antibody titration changes radically from one sample to the next, or if there's some other medical indication.
I getting a sense of deja vu, I think I've written this before.
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9 hours ago, BldBnker said:
The patient's DAT is positive, right? Has the patient received ABO incompatible platelets lately? Or received Immunoglobulin therapy (gamma globulins)? I have seen both of those scenarios cause incompatibilities with the patient's own type. Could be either Anti-A or Anti-A,B from O platelets or the gamma globulin therapy.
Good thinking. Might be interesting to see if there's anything in an eluate....
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Might be a rare IgG anti-A1 - you may not see in in the Reverse (presumably IS or buffer-only gel card), but is detected when you do anything with an antiglogulin reagent. You could try doing the Reverse by IAT. The DAT may just be a red herring, but it might represent a weird autoantibody that favors group A cells.
- Carrie Easley, Yanxia, R1R2 and 2 others
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3 hours ago, Lucky Jack said:
None of the institutions I have have worked for have added check cells to the DAT control - the check cells should be QC'd as part of daily reagent QC. From my point of view, since no AHG is added to the DAT control, the check cells serve no purpose. For negative Poly, Anti-IgG, and Anti-C3 DATs we did add check cells to verify that those reagents were not neutralized by free IgG or complement.
Please excuse my ignorance, but how can a "DAT Control" not have an antiglobulin reagent?
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On 2/3/2018 at 11:11 AM, Tabbie said:
Can anyone explain what the CD treated cell test is ? Thanks
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.
Complement QC with Poly IgG
in Transfusion Services
Posted · Edited by exlimey
Typo
My interpretation: Users of POLYSPECIFIC antiglobulin reagents are obliged to verify performance each day of use, i.e., QC should involve use of IgG-coated cells AND Complement-coated cells. This gives the user confidence that the reagent is performing as expected.
During routine testing, addition of IgG-coated cells to negative tests is sufficient to verify that the IAT was performed correctly - correct/effective washing, the antiglobulin reagent was added and is reactive, etc.
If it were a requirement to add IgG-coated cells and complement-coated cells to every negative IAT using polyspecific antiglobulin, it would be necessary to run everything in duplicate - one set would get IgG-coated cells and the other set would get complement-coated cells. I don't think that is the case.