Viable lymphocytes in frozen washed blood

Prior research has shown that cryopreservation can preserve various cell types, but the viability of lymphocytes in frozen blood remains unclear. Established methods for washing frozen blood include centrifugation and filtration, but their effectiveness varies. This gap motivated the current study to assess whether lymphocytes remain viable after freezing and washing. No prior work had resolved the optimal washing system for preserving lymphocyte function. The study aimed to address this uncertainty by comparing three washing systems. It was already known that granulocytes often show pyknotic nuclei after freezing. However, whether lymphocytes retain viability under similar conditions had not been fully established. This uncertainty drove the investigation into the effects of different washing systems on lymphocyte survival.

The aim of this study was to evaluate the viability of lymphocytes in frozen blood after washing with three different systems. The specific problem addressed was whether lymphocytes can remain functional after cryopreservation and washing. The motivation came from the need to optimize blood storage and processing methods in clinical settings. The study sought to determine if any washing system was more effective at preserving lymphocyte viability. It also aimed to assess the presence of mitotic activity in cultured lymphocytes. The researchers proposed that freezing and washing might not necessarily compromise lymphocyte function. The study focused on comparing three washing systems: Elutramatic, Haemonetics, and IBM. The findings could inform best practices for handling frozen blood samples in laboratories.

The study involved 39 samples of glycerolized, frozen, packed cells. Each sample was washed using one of three systems: Elutramatic, Haemonetics, or IBM. After washing, the cells were cultured to assess lymphocyte viability. The short-term culture method was used to detect lymphocyte growth. The samples were stored at 4°C before freezing and at -80°C for up to 40 days. The researchers examined the presence of pyknotic nuclei in granulocytes and well-preserved mononucleated cells. They also measured 3H-thymidine incorporation in PHA-induced blast cells. Mitotic figures were observed in some cultures to confirm cell division.

Out of 39 samples, 17 (43.6%) showed definite evidence of lymphocyte growth. Among these, 11 (64.7%) were washed using Elutramatic, two (11.8%) by Haemonetics, and four (23.5%) by IBM. The prefreeze and storage times were consistent across positive and negative samples. In all samples, distorted granulocytes with pyknotic nuclei were observed. Well-preserved mononucleated cells were also present. In the 17 positive samples, 3H-thymidine incorporation was detected in PHA-induced blast cells. Some cultures showed typical mitotic figures. These findings suggest that lymphocytes can remain viable after freezing and washing.

The authors propose that lymphocytes remain viable in frozen blood stored at -80°C for up to 40 days. The study found that lymphocyte growth was observed in 43.6% of samples. The Elutramatic system showed the highest success rate in preserving viable lymphocytes. The Haemonetics and IBM systems had lower success rates. The presence of mitotic figures in some cultures supports the viability of lymphocytes. The researchers suggest that the differences in lymphocyte recovery between systems require further evaluation. The findings indicate that washing methods significantly affect lymphocyte viability. These results may inform future protocols for handling frozen blood samples.