Anomalous Flotation of Separator Gel: Density or Viscosity?

Blood collection tubes containing a thixotropic polyester gel are widely used for clinical laboratory testing. The gel, because of its intermediate density (1.04 g/cm3), physically separates the liquid component of the blood from the cells (densities of 1.03 and 1.09 g/cm3 respectively) after centrifugation, and forms a barrier between the liquid phase and the cells. However, recent cases have been reported in which anomalies in the formation of this separating barrier can occur(1). Although the number of reported cases seems low, the impact on cost and the risk of reporting misleading values is substantial(1). Different interpretations have been proposed regarding whether the anomalous flotation of the gel is a matter of density or viscosity of the serum/plasma(2)(3)(4). We investigated whether this gel behavior was due to the high density or to the high viscosity of the liquid phase in a case of floating separator gel in samples from a 70-year-old woman in whom IgA λ myeloma was subsequently diagnosed. Blood samples were received in BD Vacutainer® PST™ II tubes that were processed in the routine manner. After centrifugation (1200g for 10 min) the gel was observed floating on the supernatant instead of separating the plasma from the cells. Interestingly, when BD Vacutainer SST™ II tubes were used, gel barrier formation was correct and there were no inconsistencies in the serum/clot separation. Significant laboratory data from serum included: Ca 126 mg/L, hemoglobin 81 g/L, hematocrit 24.8%, total protein 119 g/L, erythrocyte sedimentation rate 165 mm/h. Ig quantification by nephelometry showed an IgA concentration of 102 g/L, which was the highest concentration ever observed in our laboratory. These data suggested that the floating-gel phenomenon in plasma(1) was attributable to high total protein concentrations, which are associated with increases in both density and viscosity, whereas the lack of fibrinogen in serum decreased the density and/or viscosity, resulting in the correct formation of the gel barrier in the serum-separator tubes. To investigate whether the increased density or the viscosity of the plasma was responsible for the anomalous gel barrier formation, we performed experiments to simulate conditions with density and viscosity values (1.039 g/cm3 and 7.1 mPa·s, respectively) found in plasma in a case(1) with protein and Ig content quite similar to ours. We prepared solutions of dextran in solution with normal saline as solvent. Densities and viscosities were measured with an Anton Paar Density Meter-DMA 35 and an Ubbelohde-viscometer, respectively. Samples were collected into PST II tubes and centrifuged at 1200g for 10 min. Results are shown in Table 1 . Density, viscosity and position of the gel in dextran solutions and normal samples. Density, viscosity and position of the gel in dextran solutions and normal samples. Our results demonstrate that, irrespective of the high viscosity values (reference interval 1.5–2.0 mPa·s), the gel position is determined only by the density of the solution. Even in the case of the 6% dextran solution, which had a density very close to that of the gel, it was not necessary to extend the centrifugation time to get the right gel position. In summary, our results confirm that tubes containing gel separator may fail to form an appropriate barrier in blood samples with high protein content. This anomalous behavior is likely attributable to the high density of plasma/serum, because dextran solutions with very high viscosity but normal density allow proper behavior of the separator gel. These findings suggest that any substance in blood that increases density substantially can give rise to anomalous flotation of separator gel. The information provided by this study may lead to improvements in the characteristics of these gels and determination of the amount of protein that hinders appropriate gel barrier formation(1). Grant/Funding Support: None declared. Financial Disclosures: None declared.