Cryopreservation Of Blood Cells Research Articles

Establishing a method for the cryopreservation of viable peripheral blood mononuclear cells in the International Space Station

The analysis of cells frozen within the International Space Station (ISS) will provide crucial insights into the impact of the space environment on cellular functions and properties. The objective of this study was to develop a method for cryopreserving blood cells under the specific constraints of the ISS. In a ground experiment, mouse blood was directly mixed with a cryoprotectant and gradually frozen at −80 °C. Thawing the frozen blood sample resulted in the successful recovery of viable mononuclear cells when using a mixed solution of dimethylsulfoxide and hydroxyethyl starch as a cryoprotectant. In addition, we developed new freezing cases to minimize storage space utilization within the ISS freezer. Finally, we confirmed the recovery of major mononuclear immune cell subsets from the cryopreserved blood cells through a high dimensional analysis of flow cytometric data using 13 cell surface markers. Consequently, this ground study lays the foundation for the cryopreservation of viable blood cells on the ISS, enabling their analysis upon return to Earth. The application of this method in ISS studies will contribute to understanding the impact of space environments on human cells. Moreover, this method may find application in the cryopreservation of blood cells in situations where research facilities are inadequate.

Simplified Approaches for the Production of Monocyte-Derived Dendritic Cells and Study of Antigen Presentation in Bovine.

Dendritic cells are sentinels of the immune system responsible for the initiation of adaptive immune mechanisms. In that respect, the study of these cells is essential for a full understanding of host response to infectious agents and vaccines. In ruminants, the large blood volume facilitates the isolation of abundant monocytes and their derivation to other antigen-presenting cells such as dendritic cells and macrophages. However, the available protocols for the production of bovine monocyte-derived dendritic cells (moDCs) rely mostly on time-consuming and costly techniques such as density gradient centrifugation and magnetic sorting of cells. In this study, we describe a simplified protocol for the production of bovine moDC using conventional and serum-free media. We also employ moDC produced by this approach to carry out a flow cytometry-based antigen presentation assay adapted to blood fresh or frozen cells. The experimental strategies described here might enable the setup of studies involving a large number of individuals, requiring a large number of dendritic cells, or relying on the utilization of cryopreserved blood cells. These simplified protocols might contribute to the elucidation of cell-mediated immune responses in bovine.

5. “What can we learn from cryopreserved blood cells?”

5. “What can we learn from cryopreserved blood cells?”

Complex Karyotype in Patients with Chronic Lymphocytic Leukemia.

Chronic Lymphocytic Leukemia (CLL) is heterogeneous and there are two clinical forms with different outcome, in which initial stage and parameters of tumor burden, expression of CD38 and /or ZAP 70, chromosome abnormalities such as 11q23 and 17p13 are associated with a more aggressive course of disease.The aim of this study is to analyse clinical and biological characteristics and outcome of patients with complex karyotype, and to asses if there are differences with other genomic aberrations.Fifty patients diagnosed of CLL between 1980 and 1994 were studied (median age: 71 yrs; female/male: 18/32; median follow up 6.6 yrs.). Biological studies with flow cytometry and fluorescence in situ hybridization were done on cryopreserved blood cells. Clinical stages at diagnosis were: Rai stage 0: 56%; I: 20%; II: 16%; III: 6%; IV: 2%. Binet A: 72%; B: 20%; C: 8%. At the time of diagnosis 37% of patients needed treatment and 61% presented disease progression. At the time of this study, 10% of patients are alive with a median follow up of 15.3 yrs.Seven patients (28%) presented complex karyotype. At the time of diagnosis 43% patients needed treatment and clinical stages were: Rai stage O: 29%, I: 14%, II: 43%, III: 14%, IV: 0%. Binet A: 43%; B: 43%; C: 14%. Median overall median survival was 72 months.The most frequent genomic aberrations associated with complex karyotype were: del (17p13) and trisomy 12 (60 %). Cellular expression of CD38 and ZAP 70 was not able to separate this group.Compared with other chromosome abnormalities, there were not important differences in clinical stage at diagnosis or expression of CD38 or ZAP 70, but overall median survival could distinguish three groups: del 11q22-23 or del 17p13 (34 months, p < 0,05) complex karyotype (72 months), and trisomy 12 or del 13q14 (79 months, p > 0,05).In conclusion, complex karyotype is frequently formed by del 17p13 and trisomy 12 and identifies a subgroup of patients with better prognosis compared with isolated p53 deletion. More studies have to be realized to determine if trisomy 12 is a cell defence mechanism.

Cryopreserved chronic lymphocytic leukemia cells analyzed by multicolor fluorescence in situ hybridization after optimized mitogen stimulation.

We investigated the utility of multicolor in situ fluorescence hybridization (mFISH) on cryopreserved blood cells from 11 chronic lymphocytic leukemia (CLL) patients. The results demonstrate that an individually chosen optimized mitogen combination induces proliferation of neoplastic B-cells after cryopreservation. Abnormal cells were detected in eight samples by mFISH, and, in six samples, the abnormality could be verified by comparative genomic hybridization or interphase FISH. In addition to typical CLL abnormalities, such as del(11q) or +12, several balanced translocations and single-cell abnormalities were found. Thus, mFISH can reveal new prognostically relevant chromosome aberrations in CLL.

An apparatus for regulating the freezing and thawing rates in the cryopreservation of blood cells

An apparatus is described which was developed to freeze small quantities of liquid at rates of up to 60 °C per sec and to thaw them at up to 500 °C per sec. The apparatus is especially suited to systematically ascertaining the optimum freezing and thawing rates and their effects on the viability of blood cells in the course of cryopreservation. The sample container is a metal tube cooled with liquid nitrogen and heated by an electric current. The temperature of the sample is sensed by a thermocouple and controlled by an electronic control device via the amount of heater current. The proper knowledge of the optimum preservation conditions provides satisfactory conditions for the development of processes for the cryopreservation of sufficiently large quantities of blood cells for transfusion purposes.