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Antigen typing in the Gel System


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Is there anyone who can tell me if you can do your antigen typing on the gel cards? I know that they sell the RBC Phenotype cards, and the C, c, e, and E, but what about the others such as K, Jka, Jkb, etc? Does anyone have a method of performing these other than tubes?:confused:

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We currently perform all of our antiglobulin antigen typing using the IgG gel cards. We use 50ul of 0.8 cells and 25ul of the antisera, we have been doing this for 8 years now with no problems. We are currently validating the monoclonals using buffered gel cards. So far so good. The only ones that we will continue do in tube will be M and N.

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We do almost all of our antigen testing via DiaMed gel cards, have done so for a good five to ten years now and have experienced no problems whatsoever.

Obviously, you have to validate each of your antisera before use, and then validate each new batch, but as you would be running positive and negative controls with each batch of tests you are performing, this is not as onerous as it sounds.

We use the same method as swede.

:):):)

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We use gel for typing our donors for Duffy, Kid , S, and s(basically all antiglobulin reagent). and use same method as above. 50ul of donor cell suspension(in MTS 2) plus 25ul of antisera...it's saves us lots of reagent and money.

Is anyone out there using ProVue?

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We use gel for typing our donors for Duffy, Kid , S, and s(basically all antiglobulin reagent). and use same method as above. 50ul of donor cell suspension(in MTS 2) plus 25ul of antisera...it's saves us lots of reagent and money.

Is anyone out there using ProVue?

We are currently working out the contract to get the Provue.....we are hoping to get it up and running by the end of the year. Are you using ProVue? If so, how do you like it?

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We are currently working out the contract to get the Provue.....we are hoping to get it up and running by the end of the year. Are you using ProVue? If so, how do you like it?

We have been using ProVUe for almost 5 years and I love it. If ProVUe is down, my techs can not handle the workload anymore because they are so use to ProVue now.

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No our workload has gone up and started doing lots of reference work in house. We do not use ALBA QC as it is outregously expensive. I couldn't justify the cost. We are preparing our own using patient specimen and COnfidence kit.

I do not have that many down time..sometime it is tech fault(forgot to remopve cap), though it's very rare.

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We also just got done with all of our validation for gel ag typing, so far so good. We're using the same procedure as Swede's posting. We're using the MTS Rh phenotyping card and monoclonal C/E/c/e cards, and the IAT phase antisera with IgG cards for the rest fo the major antigens.

Our only observation thus far is to beware of "hazy" reactions in the gel column. You'll find one example of a hazy column in Ortho's interpretation guide; usually seen with high levels of plasma proteins. We washed the cells and repeated testing and saw less haziness, but it didn't go away completely. When in doubt, you can always confirm in tube. Just make sure that techs don't interpret this as a positive result without further investigation - both of our cases were indeed negative for the ag being tested.

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We also just got done with all of our validation for gel ag typing, so far so good. We're using the same procedure as Swede's posting. We're using the MTS Rh phenotyping card and monoclonal C/E/c/e cards, and the IAT phase antisera with IgG cards for the rest fo the major antigens.

Our only observation thus far is to beware of "hazy" reactions in the gel column. You'll find one example of a hazy column in Ortho's interpretation guide; usually seen with high levels of plasma proteins. We washed the cells and repeated testing and saw less haziness, but it didn't go away completely. When in doubt, you can always confirm in tube. Just make sure that techs don't interpret this as a positive result without further investigation - both of our cases were indeed negative for the ag being tested.

Agreed, but also beware of ignoring weak reactions with the anti-e, without a thorough investigation, particularly if the patient is from the Black population; it may be a case of CdeS.

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Good point! We will also further investigate any weak reactivity - maybe molecular testing as well. All of our typings so far are much stronger in gel (at least 2+, usually 4+) and almost exclusively on our sickle cell patients. (lots of variant antigens)

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Hi all,

Interesting post to me as we are validating tube reagents in BioVue and DiaMed ID-MTS cards.

We can get most tube reagents to perform well except for M, N, S, Lea and Leb.

For those who are attempting this or using it poutinely I offer a few points to consider.

1. M and N phenotyping reagents we have tried often display unusual specificity. Anti-N for example is almost always an anti-glycophorin A and will therefore detect all human red cells. If you dilute it enough it will have apparent specificity for N and work fine BUT if you change the test sensitivity, you can get into a tangle. So when working up the volumes and perhaps the dilutions used, be very careful and precise and validate it carefully. Also be aware that a change in reagent batch (for example if a new batch is a little stronger) can change the specificy. It may still work fine in tube but start detecting N- cells in CAT, so be careful of batched and perform preacceptance testing when you change reagent batches.

2. Anti-M seems to show similar effects.

3. I have my doubts that tube Le phenotyping reagents are usable (or perhaps should be used) in CAT. As the strengths of Le antigens vary so much and are dependant on many other attributes like ABO group of the cells and ethnicity. For example in a normal group A individual the amount of Leb antigen made is much less than in a group O as much of the Leb in a group A exists as ALeb antigen - an antigen that does not react with anti-Leb. Therefore the level of Leb is always reduced in group A individuals and it often shows variable reactivity and can frequently type as Le(a-b-) frequently in Europeans (Larson et al 1996). The Le(a+b+) phenotype, which is the second most frequent phenotype in Asians, Polynesians and Aborigines is caused by a "weak" secretor gene. To be an Le(a+b+) phenotype a person must have both weak expression of Lea and weak Leb. As a consequence, they are often incorrectly phenotyped with commercial reagents that have their Le antigen testing sensitivity that is "fine tuned" by manufacturers based on reactions seen in Caucasian Lewis phenotypes. When the Le(a+b+) phenotype is further compounded with the weak expression of Leb seen in a group A then the ability to determine an accurate phenotype become virtually impossible. Genetic studies has shown that the Le(a+b+) phenotype is usually incorrectly phenotyped, and much more so in group A. Only genotyping can accurately confirm this status.

Here are some referrences below if you are interested:

Henry S, Mollicone R, Fernandez P, Samuelsson B, Oriol R, Larson G: Molecular basis for erythrocyte Le(a+b+) and the salivary ABH partial-secretor phenotypes. Expression of a FUT2 secretor allele with an A→T mutation at nucleotide 385 correlates with reduced α(1,2)fucosyltransferase activity. Glycoconj J 1996; 13:985 993.

Henry S. Le(a+b+); phenotype and genotype. NZ J Med Lab Sci 1996;50:50 53

Henry S, Mollicone R, Fernandez P, Samuelsson B, Oriol R, Larson G: Homozygous expression of a missense mutation at nucleotide 385 in the FUT2 gene associates with the Le(a+b+) partial-secretor phenotype in an Indonesian family. Biochem Biophys Res Commun 1996;219:675 678.

Henry SM: Review: phenotyping for Lewis and Secretor histo-blood group antigens. Immunohem 1996; 12:51 61

Henry SM, Oriol R, Samuelsson BE: Lewis histo-blood group system and associated secretory phenotypes. Vox Sang 1995;69:166 182.

Henry SM, Simpson LA, Benny AG, Woodfield DG: Investigation of Polynesian Lewis phenotypes. Variability in detection of Lewis antigens by monoclonal, goat and human antisera. NZ J Med Lab Technol 1989;43:64 67.

But then again maybe lewis phenotyping inaccuracies simply do not matter!!!!!

I would be interested in anybodies experiences with the use of tube based M, N, S and Lea and Leb in CAT where you think you have good performance.

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Is there anyone who can tell me if you can do your antigen typing on the gel cards? I know that they sell the RBC Phenotype cards, and the C, c, e, and E, but what about the others such as K, Jka, Jkb, etc? Does anyone have a method of performing these other than tubes?:confused:

You can identify the other antigens by single antigen or entigen profiles by DiaMed Microtyping System.

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