San Diego VA HCS San Diego, California, United States
Background/Case Studies: Determine if manual gel (gel) could replace tube phenotyping for Fya in our blood bank. Wanted: fast, reliable, robust system that did not require cell washers, glass tubes, AHG phase reactions, incubation. Want to avoid antigen negative units from blood suppliers (expensive and incurs time delays). We already use gel as backup to an automated testing system (Ortho Vision). Wanted to see if antisera from different companies could be used in the system. If successful will validate other RBC antigen antisera.
Study
Design/Methods: Tube testing was done according to SOP. Gel was done with Ortho Workstation. All reagents passed QC. We used Immucor anti-Fya antisera (25uL) with Ortho 0.8% Panel A cells (50uL) in IgG cards. Fya phenotyping requires 15" at 37C so gel cards were also incubated 15" at 37C. Duplicate reactions were run in gel minus incubation to see if it could be omitted. Gel cards were spun for 10". Next I tested 12 RBC units and 6 untransfused patient samples in tube, then in gel, both with 15" 37C incubation and without incubation. Donor RBC and patient samples were washed 1X with saline (tube) or Ortho Diluent (gel), resuspended to 3% (tube), or 0.8% (gel). These were tested with Ortho anti-Fya antisera without changes. Last anti-Fyb, anti-K, and anti-Jkb (Immucor) were tested in tube and gel as before, but with 15" incubation at RT, and Buffer gel cards. Duplicate reactions were set up without incubation.
Results/Findings: Gel phenotyping outperformed tube method for antisera tested. Reactions were always stronger with no false positives or negatives. Same gel reaction strength was seen without incubation. Donor units and patients could be tested. Donor units were collected in different solutions which did not affect testing. Patient samples (3 days-2 weeks old) all tested correctly compared to tube. No questionable results for any gel testing occurred. (Table 1). Conclusions: Gel has advantages over tube method. Less reagent, less waste, no glassware, no cell washer No ordering typed units from blood supplier. Gel cards can be stored room temperature. Gel could be used to phenotype various RBC antigens. Gel reactions can be saved by taping the card for review. The gel centrifuge step is only 10min and other testing can be done concurrently. Gel requires one simple machine and pipettes. Less chance of false negatives (more robust reactions) and no false positives. Tube and man gel agreed 100%. Important for antigenically weak donors. IgG and Buffer gel cards are needed but costly specific antisera cards are not required. Similar typing reagents from different suppliers worked equally well. Elimination of the 15" incubation saves time when antigen typed units are urgently needed and could prevent transfusion of untyped RBC units in patients with dangerous antibodies. The gel system is simple, works as an adjunct to other phenotyping methods or can stand alone in a hospital blood bank.
Importance of research: Considerable time and money can be saved with this method. No need to incubate reactions for any of the antisera that were tested. Less waste, no glass waste, no need for special antigen cards, no need for cell washer. Easy scalable system. Less technique dependent. All results are in 100% complete agreement compared to standard tube methodology. Reactions are more robust. Less reagent needed. Removes danger of false negatives without the chance of false positives. Highly efficient safe system.
