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Antibody products must be evaluated on relevant tissues, which can often be rare, difficult, and expensive to obtain. Cell lines can offer an optimal solution, and so a method to mimic frozen tissue blocks with cryoembedded cultured cell pellets was developed. Kidney sections from a patient with systemic lupus erythematosus (SLE) in the flare state were compared to cultured, cryoembedded Raji cells by detecting the complement components C1q, C3, and C4 on slides from both. An immunocytochemical (ICC) assay to detect C1q, C3, and C4 immune complexes on the Raji cell membranes was developed. The slides from the cryoembedded Raji cells were acceptable compared to the SLE kidney tissue for 99% of the C1q staining, 90% of the C3 staining, and 91% of the C4 staining. The method was also tested with a mixture of Raji cells and SW480 colorectal adenocarcinoma cells, and found that antibody staining the complement components was acceptable in the cell mixture and that the SW480 cells had acceptable staining with anti-MSH2 antibody. Further, BT-474 breast cancer cells were stained and prepared by a cryoembedding method with an anti-HER-2/neu antibody and, again, acceptable staining was observed. The method preserves cellular morphology, antigenicity, and cell-to-cell contact while avoiding freeze artifacts and the issues that arise from fixing cells, such as antigen masking and auto-fluorescence. The method to prepare cryoembedded tissue blocks that mimic frozen tissue blocks is widely applicable to the field of cytopathology. [ABSTRACT FROM AUTHOR]

Objective: To establish reliable methods for cryopreservation of human embryonic stem cells (hESCs). Design: Prospective experimental study. Setting: University laboratory. Patient(s): One hESC line. Intervention(s): The attachment rates and recovery rates of cryopreserved hESCs using three different cryopreservation methods were compared. Main Outcome Measure(s): The hESCs were frozen and thawed by conventional cryopreservation, programmable cryopreservation, and vitrification method. The efficiency of cryopreservation was assessed by attachment rate and recovery rate. Result(s): The attachment rate and recovery rate after thawing of hESCs frozen by the conventional cryopreservation method were significantly lower than those of hESCs frozen by programmed cryopreservation and vitrification methods. Vitrification resulted in the highest attachment rate and recovery rate compared with the other two methods. Human ESCs after vitrification and programmable cryopreservation still expressed pluripotent markers, maintained normal karyotype, and retained their pluripotency. Conclusion(s): Our data show that programmable cryopreservation and vitrification methods are appropriate for cryopreservation of hESCs, whereas the conventional slow-rate freezing method is not appropriate for cryopreservation of hESCs. [Copyright &y& Elsevier]

To evaluate the ability of two cryopreservation methods and three cryoprotectants to preserve sperm quality.A prospective clinical study.Male infertility clinic at a tertiary healthcare center.Twenty infertile men and 10 healthy donors.In the first experiment, semen was cryopreserved by either the Irvine Scientific method (IS) or the Cleveland Clinic Foundation (CCF) method. In the second experiment, semen was cryopreserved by the IS method and one of three cryoprotectants: TES and Tris yolk buffer, Sperm Freezing Medium, or Enhance Sperm Freeze.Postthaw sperm motility, cryosurvival, and kinematics.Percentages of postthaw sperm motility and cryosurvival were higher in the IS cryopreservation method compared with in the CCF method (15.94 ± 9.19 vs. 12.07 ± 7.31 and 47.42 ± 17.44 vs. 35.76 ± 17.56). However, the CCF method resulted in significantly better sperm kinematics. Postthaw motility in the donors and patients was highest in the samples frozen in TES and Tris yolk buffer medium.The IS method was associated with more flash freezing compared with the CCF method and resulted in better preservation of sperm motility and a higher cryosurvival rate. TES and Tris yolk buffer was most effective at protecting sperm from the negative effects of the cryopreservation process. This may be due to the presence of egg yolk along with glycerol. [Copyright &y& Elsevier]