Comparison of conventional tube and gel-based agglutination tests for feline AB system blood typing

Gel technology is widely used in human medicine for blood typing. It carries many advantages over routine tube testing, such as: standardization, stability, smaller sample volume, easy to perform and read, and rapidity. The aim of this study is to evaluate the gel column technique in feline blood typing. The blood type of one hundred and thirty-six blood samples anticoagulated with EDTA or CPDA from feline blood donors, feline blood recipients, health patients and stored units of whole blood was determined using tube agglutination (TUBE) with plasma from type B cats as anti-A reagent, Triticum vulgaris lectin as anti-B reagent and PBS for control. Samples positive for type B and AB were back typed with type A RBCs to confirm whether the samples were B (strong agglutination) or AB (absence of agglutination). Samples were blood typed in duplicate using the same anti-A and anti-B reagents in a neutral gel (GEL) column technique (ID-Card NaCl enzyme test and cold agglutinins, DiaMed). Briefly, 25 μL of type B plasma and 25 μL of Triticum vulgaris lectin were mixed with 50 μL of a 0.8% RBC suspension (made by suspending 10 μL of the RBC pellet in 1 mL of low ionic strength solution) in the reaction chamber of a gel column identified as A and B respectively. For all samples, a negative control column containing the RBC suspension of interest and PBS was included. The gel columns were incubated for 15 min at room temperature and then centrifuged in a special gel column card centrifuge (ID-Centrifuge 24 S, DiaMed) at 80 g for 10 min. Finally, the gel column cards were visually checked to identify positive samples via agglutination reactions. Results were considered valid if the control column was negative. Sensitivity (Se), specificity (Sp), positive predictive value (PPV) and negative predictive value (NPV) and Cohen’s kappa coefficient (K) for GEL were calculated, considering TUBE as the gold standard technique. Of 136 samples typed with TUBE, 95 (69.8%) were type A, 22 (16.2%,) type B and 19 (14.0%) type AB. All B and AB samples were confirmed by back typing. With GEL 112 samples (82.3%) gave concordant results with TUBE, and 24 samples showed a mixed-field agglutination pattern (presence of a layer of RBCs simultaneously either at the top and at the bottom of the gel in A or in B gel column). If a mixed-field pattern was interpreted as a negative result 135/136 (99.3%) samples showed concordant results and Se, Sp, PPV and NPV (95%CI) were respectively 100% (96.1-100), 100% (91.4-100), 100%, 100% for type A, 95.4% (77.1-99.8), 100% (96.8-100), 100% and 99.1% (94.3-99.8) for type B, 100% (82.3-100), 99.1% (95.3-99.9), 95.0% (72.9-99.2) and 100% for type AB. Strength of agreement was very good (K= 0.98, 95%CI 0.95-1.00). The GEL column technique, using the same anti-A and anti-B reagents as in TUBE test is a sensitive and specific method for blood typing feline samples. Mixed-field pattern should be considered as negative results.