Evaluation of post-transfusion RBC alloimmunization in dogs using a gel-column crossmatch with and without anti-canine globulin enhancement.

A blood crossmatch is essential to ensure RBC compatibility for previously transfused dogs. There is no gold standard crossmatch method for dogs, although the standards used most commonly by academic institutions and reference laboratories are the tube and gel-column crossmatches. Addition of anti-canine globulin (ACG) has been suggested to increase detection of RBC incompatibilities. Our objective was to determine if there is a correlation between results of a standard and an ACG-enhanced gel-column crossmatch in detecting post-transfusion RBC alloimmunization. Pre- and post-transfusion serum or plasma samples were obtained from 33 dogs for major crossmatches to 1-6 (median: 3) blood donors. Crossmatches were performed with (n = 202) and without (n = 202) ACG, with results scored by 4 observers, 3 of whom were anonymized. Ten of 33 (30%) dogs had major crossmatch incompatibilities post-transfusion. RBC incompatibilities (2-4+ agglutination) were detected only with ACG in 4 dogs, only without ACG in 3 dogs, and with both methods in 3 dogs. There was fair correlation between crossmatch methods for determination of compatibility (ρ = 0.34; p < 0.001) and incompatibility (ρ = 0.35; p < 0.001) scores. Among 4 observers, there was near-perfect agreement in determining compatibility (κ = 0.97; p < 0.001) and substantial agreement in overall scoring of incompatibility (κ = 0.77; p < 0.001). Our results suggest that detection of RBC incompatibilities in dogs can be maximized by performing a gel-column crossmatch both with and without ACG enhancement.

Effect of donor blood storage on gel column crossmatch in dogs.

BACKGROUND: Compared with fresh blood, stored equine donor blood results in spurious tube crossmatch incompatibilities. Interpretation of blood crossmatch results is considered subjective. OBJECTIVES: We aimed to determine if the duration of canine donor blood storage impacts compatibility testing using a standard gel column crossmatch and evaluate interobserver variation in the interpretation of crossmatch results. METHODS: Observational study. Whole blood segments were obtained from 23 canine packed red blood cell (RBC) units for use in crossmatches after storage for 0, 7, 14, 21, 28, and 35 days. Major and minor crossmatches were performed using serum and RBCs, respectively, from two to three healthy "recipient" dogs per unit. All crossmatch results were interpreted by four observers, of whom three were blinded. RESULTS: All major crossmatches (n = 61) were compatible on day 0 and remained compatible through day 35 of storage. All minor crossmatches (n = 69) were compatible at all time points, except for five donor pairs with 1 to 3+ agglutination. Repeat testing of these five donor pairs confirmed crossmatch incompatibilities on days 0 through 35, with no change in the degree of incompatibility over time. There was substantial agreement among four observers in determining compatibility (κ = 0.94) and scoring incompatibility (κ = 0.76). CONCLUSIONS: The current practice of performing canine crossmatches with whole blood segments stored for up to 35 days is acceptable, with no spurious changes in compatibility expected over time. The substantial interobserver agreement suggests that the gel column is suitable for performing canine crossmatches in a laboratory setting with multiple personnel.

Hemolytic anemia blunts the cytokine response to transfusion of older red blood cells in mice and dogs.

BACKGROUND: Transfusion of red blood cells (RBCs) stored for longer durations induces hemolysis and inflammatory cytokine production in murine and canine models. Despite immune system activation by stored RBCs, human randomized trials suggest that fresher RBC transfusions do not improve clinical outcomes. We hypothesized that underlying recipient hemolysis may affect cytokine responses to older RBC transfusions. STUDY DESIGN AND METHODS: C57BL/6 mouse cohorts were infused with anti-TER119 antibody to induce hemolysis, rabbit anti-platelet antiserum to induce immune thrombocytopenia (ITP), or appropriate control antibodies. Two days later, mice were transfused with fresh or stored RBCs. Furthermore, in a prospective, randomized, blinded trial, 38 client-owned dogs with primary autoimmune hemolytic anemia (AIHA) and two dogs with ITP, requiring RBC transfusion, were enrolled and randomized to receive fresh (≤7 days) or old (≥21 days) stored RBC transfusions. Monocyte chemoattractant protein (MCP)-1 levels were assessed at defined times after transfusion. RESULTS: Prior immune-mediated hemolysis blunted the MCP-1 response to stored RBC transfusion in mice (361 ± 111 pg/ml vs. 6836 ± 1528 pg/ml in mice with immune hemolysis vs. ITP, respectively; mean ± SD; p < .0001). Although hemolysis markers increased after transfusion of older RBCs, the cytokine response was also muted in dogs with AIHA. No differences in morbidity or mortality were evident comparing dogs randomized to fresh or old RBCs. CONCLUSION: These data suggest that underlying hemolysis blunts inflammatory responses to old RBC transfusions. The canine data support randomized trial results suggesting a lack of clinical benefit with fresh RBC transfusions in subjects with underlying, baseline hemolysis.

Influence of canine donor plasma hemostatic protein concentration on quality of cryoprecipitate.

BACKGROUND: Cryoprecipitate (CRYO) is a plasma component containing high concentrations of factor VIII (FVIII), von Willebrand factor (VWF), and fibrinogen. Because Greyhounds are reported to have lower plasma VWF and fibrinogen concentrations, their plasma may not yield high potency CRYO. OBJECTIVES: To determine if plasma hemostatic protein concentration is a good predictor of CRYO potency and if a difference exists in quality of CRYO prepared from Greyhounds versus non-Greyhounds. ANIMALS: Twenty Greyhounds and 20 non-Greyhounds. METHODS: A 450 mL unit of blood was collected from each donor, centrifuged to prepare fresh frozen plasma (FFP), and processed to CRYO. Aliquots of FFP and CRYO were analyzed for FVIII, VWF, and fibrinogen content and factor recovery. RESULTS: A positive correlation was found among donor plasma FVIII, VWF and fibrinogen concentration, and CRYO factor content (P < .001). Mean recovery was highest for VWF (67%), followed by fibrinogen (47%), and FVIII (37%). No breed difference was found in mean CRYO FVIII content, but CRYO VWF and fibrinogen were lower in Greyhounds (P = .004 and P < .001, respectively). No difference was found between Greyhounds and non-Greyhounds for the number of CRYO units meeting human blood banking standards. CONCLUSIONS AND CLINICAL IMPORTANCE: Factor concentration in FFP is associated with CRYO potency, suggesting that prescreening of blood donors may enhance CRYO quality. Despite lower VWF and fibrinogen content, CRYO prepared from Greyhounds is acceptable based on blood banking standards for humans, indicating that Greyhound FFP does not need to be excluded from CRYO production.

Prevalence of naturally occurring non-AB blood type incompatibilities in cats and influence of crossmatch on transfusion outcomes.

BACKGROUND: Recognition of the feline red blood cell (RBC) antigen Mik and the presence of naturally occurring anti-Mik antibodies resulting in acute hemolytic transfusion reactions prompted the recommendation to perform a crossmatch before a cat's first RBC transfusion, but this guideline has not yet become a standard practice. OBJECTIVE: To determine the prevalence of naturally occurring non-AB alloantibodies detectable by tube crossmatch, and to compare transfusion outcomes in cats with and without a crossmatch performed. ANIMALS: Three hundred cats that received an RBC transfusion, with or without a major crossmatch performed. METHODS: Retrospective study. RESULTS: Major crossmatch incompatibilities were documented in 23 of 154 transfusion-naive cats (14.9%) and in 15 of 55 previously transfused cats (27%; P = 0.042). Type-specific packed RBCs (pRBCs) were administered to 167 and 82 cats with and without a crossmatch, respectively. Median volume of pRBCs administered during the first transfusion was 5.3 mL/kg (range, 2.4-18 mL/kg). Median change in PCV scaled to dose of pRBCs was +0.8%/mL/kg; administration of crossmatch-compatible pRBCs was not associated with a greater increase in PCV. Febrile transfusion reactions occurred more often in cats that received non-crossmatched (10.1%) compared to crossmatched (2.5%) pRBCs (P = 0.022). Seventy-six percent of cats that received pRBC transfusions survived to hospital discharge. A crossmatch was not associated with improved survival to discharge or at 30 or 60 days posttransfusion. CONCLUSIONS AND CLINICAL IMPORTANCE: The prevalence of naturally occurring non-AB incompatibilities is sufficiently high to justify the recommendation to perform a crossmatch before all (including the first) RBC transfusions in cats.

Evaluation of the use of shock index in identifying acute blood loss in healthy blood donor dogs.

OBJECTIVE: To determine if shock index (SI) would increase following blood donation and if it would be a more sensitive assessment of acute blood loss as compared with heart rate (HR), blood pressure, and plasma lactate. DESIGN: Prospective study. SETTING: University teaching hospital. ANIMALS: Twenty client-owned clinically normal dogs. INTERVENTIONS: Peripheral venous blood measurements and blood donation. MEASUREMENTS AND MAIN RESULTS: Data were collected at 3 time points: prior to donation (Tpre ), immediately after donation (T0 ), and 10 minutes following completion of donation (T10 ). HR and systolic blood pressure (SBP) were recorded and used to calculate SI at time points Tpre , T0 , and T10 . Packed cell volume (PCV), total plasma protein (TPP), and plasma lactate were evaluated from a peripheral venous blood sample at Tpre and T10. The mean SI was significantly increased at both time points following blood donation as compared to baseline (SIpre = 0.88 ± 0.19 vs SI0 = 1.17 ± 0.21 vs SI10 = 1.12 ± 0.25 (P = 0.0002 and 0.0003, respectively). Following blood donation, the mean SBP was significantly lower (SBPpre = 149 ± 24 mm Hg, SBP0 = 118 ± 20 mm Hg; P = 0.0001, SBP10 = 133 ± 21 mm Hg; P = 0.011). The mean HR was not significantly different at T0 but was significantly increased at T10 (HRpre = 128 ± 21/min, HR0 = 136 ± 25/min, P = 0.193; HR10 = 146 ± 29/min, P = 0.003). There was no significant difference in mean PCV (PCVpre = 50 ± 4%, PCV10 = 48 ± 4%, P = 0.08). The mean TPP and plasma lactate were significantly different following donation but still within the reference interval (TPPpre = 6.8 ± 0.4 g/dL, TPP10 = 6.4 ± 0.4 g/dL, P = 0.0014; Lacpre = 1.7 ± 0.7mmol/L, Lac10 = 1.9 ± 0.8 mmol/L, P = 0.04). A receiver operating characteristic (ROC) analysis comparing area under the curve (AUC) for SI, HR, and SBP at T0 and T10 compared to Tpre found that SI (AUC at T0 : 0.858, CI: 0.730, 0.984 AUC at T10 : 0.769 CI: 0.617, 0.921) was a better indicator of blood loss than SBP at both T0 (AUC 0.165, CI: 0.0384, 0.292, P < 0.0001) and T10 (AUC 0.288, CI: 0.124, 0.451, P < 0.001) and better than HR at T0 (AUC 0.574, CI: 0.392, 0.756, P < 0.001). An SI cut-off of 1.064 was 80% specific and 85% sensitive at T0 for detecting blood loss. CONCLUSIONS: An SI > 1.0 is a sensitive and specific tool for detecting acute small volume blood loss in healthy dogs.