Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “cryopreservation”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources
PubMed2026

Cryopreservation-induced proteomic alterations in Pêga donkey (Equus asinus) spermatozoa.

Semen cryopreservation is a key tool for assisted reproduction and genetic conservation, but its efficiency remains limited in donkeys, compromising post-thaw sperm quality. This study aimed to characterize the proteomic profile of Pêga donkey spermatozoa and to investigate molecular mechanisms associated with cryopreservation-induced impairment of sperm function. Semen samples were collected from Pêga jacks and evaluated for sperm motility and vigor before and after cryopreservation. Quantitative proteomic analysis was performed by LC-MS/MS, followed by bioinformatic characterization of differentially abundant proteins. Cryopreservation markedly reduced sperm motility in all animals, whereas sperm vigor showed only a non-significant tendency toward reduction, suggesting sublethal cryoinjury primarily affecting flagellar efficiency. Proteomic profiling identified 554 proteins, of which 98 were differentially abundant between in natura and cryopreserved spermatozoa. Functional enrichment analyses showed that these proteins were mainly associated with energy metabolism, mitochondrial oxidative phosphorylation, glycolysis, cytoskeletal organization, signal transduction, proteostasis, and oxidative stress response. Notably, proteins involved in ATP production, mitochondrial function, and axonemal organization were significantly altered, supporting a mechanistic link between metabolic dysfunction, flagellar structural disorganization, and reduced post-thaw motility. Overall, cryopreservation induced coordinated and compartment-specific remodeling of the donkey sperm proteome, particularly affecting pathways essential for motility and functional competence. These findings provide new molecular insights into the cryobiological vulnerability of donkey spermatozoa and establish a mechanistic basis for the development of more effective, biology-driven cryopreservation strategies for this species.

Animals↗
PubMed2026

Research note: Efficient preservation of genome-edited chicken germplasm via N-methylacetamide-based semen cryopreservation.

The rapid development of genome-edited chicken lines for agricultural and biomedical use requires effective methods for long-term preservation. In birds, cryopreservation of oocytes and embryos is challenging due to the structure of the egg. This makes semen cryopreservation one of the most practical alternatives, however, this method has not been validated in genome-edited chickens. This study evaluated the feasibility of cryopreserving semen from genome-edited chickens using an N-methylacetamide (N-MA)-based protocol. Two genome-edited chicken lines were used in this study which are a DAZL-GFP germ cell reporter line and a RAG1 knockout line. Semen was stored for either one week or one month prior to assessing fertility and hatchability. The results demonstrated that cryopreserved semen from both genome-edited chicken lines maintained their fertilizing capacity, with no significant differences in fertility or hatchability compared to cryopreserved wild-type groups at either storage duration. Hatchability remained above 80% across all groups, indicating that semen cryopreservation did not significantly affect embryo development after successful fertilization. Furthermore, SYBR-14/PI staining revealed no significant differences in sperm viability between genome-edited and control groups, although both showed a significant decrease in viability compared to fresh semen. These findings demonstrate that N-MA-based semen cryopreservation provides a reliable and practical method for the preservation of genome-edited chicken lines.

Chicken↗
PubMed2025

CryoSCAPE: Scalable immune profiling using cryopreserved whole blood for multi-omic single cell and functional assays.

BACKGROUND: The field of single cell technologies has rapidly advanced our comprehension of the human immune system, offering unprecedented insights into cellular heterogeneity and immune function. While cryopreserved peripheral blood mononuclear cell (PBMC) samples enable deep characterization of immune cells, challenges in clinical isolation and preservation limit their application in underserved communities with limited access to research facilities. We present CryoSCAPE (Cryopreservation for Scalable Cellular And Proteomic Exploration), a scalable method for immune studies of human PBMC with multi-omic single cell assays using direct cryopreservation of whole blood. RESULTS: Comparative analyses of matched human PBMC from cryopreserved whole blood and density gradient isolation demonstrate the efficacy of this methodology in capturing cell proportions and molecular features. The method was then optimized and verified for high sample throughput using fixed single cell RNA sequencing and liquid handling automation with a single batch of 60 cryopreserved whole blood samples. Additionally, cryopreserved whole blood was demonstrated to be compatible with functional assays, enabling this sample preservation method for clinical research. CONCLUSIONS: The CryoSCAPE method, optimized for scalability and cost-effectiveness, allows for high-throughput single cell RNA sequencing and functional assays while minimizing sample handling challenges. Utilization of this method in the clinic has the potential to democratize access to single-cell assays and enhance our understanding of immune function across diverse populations.

Humans↗
PubMed2026

Protective effects of seminal exosomes on cryopreserved sperm via inhibiting oxidative damage.

This study aimed to explore the protective effect of seminal plasma exosomes (SPEs) on human sperm structure and function during cryopreservation and its potential mechanism. The samples were divided into two groups: the control group was treated solely with sperm cryoprotectant before freezing, while the exosome group was supplemented with SPEs. After cryopreservation and thawing, sperm progressive motility, normal morphological rate, and survival rate were evaluated. Furthermore, PKH67 labeling experiments were performed, and oxidative stress markers as well as energy metabolism indicators in sperm were detected. Subsequent mechanism exploration was conducted via proteomic analysis and protein validation assays. This work reveals that adding SPEs at a concentration of 1 or 2 mg/ml effectively improves sperm progressive motility after cryopreservation. After supplementing with SPEs, sperm glucose levels are reduced and mitochondrial membrane potential is enhanced. Simultaneously, SPEs alleviate oxidative stress by decreasing reactive oxygen species (ROS) and DNA fragment index (DFI) while increasing superoxide dismutase (SOD) activity. Functional annotation of proteomics reveals that 14 of the differentially expressed proteins (DEPs) are associated with sperm motility. Enriched metabolic pathways related to sperm motility and sperm protein validation experiments indicate that the expression of MAPK, p-MAPK, and p-JNK proteins in sperm is higher in the Exosome group than in the Control group. This study provides important theoretical support for the application of SPEs in mitigating cryopreservation damage to sperm by enhancing antioxidant capacity. The specific mechanism may be mediated by the MAPK/p-JNK pathway.

Male↗
PubMed1978

Advantages of cryopreserved lymphocytes for sequential evaluation of human immune competence. II. Mixed lymphocyte cultures and mononuclear cell subpopulations.

Peripheral blood lymphocytes were obtained from normal volunteer donors over a 2-week period. One-way or two-way mixed lymphocyte reactions, percentages of T-cells and B-cells, and percentages of monocytes were quantitated on both fresh and cryopreserved lymphocytes. These comparisons revealed that the measurement of mixed leukocyte reactions, fractions of E-rosettes and EA rosettes, and the fraction of esterase-staining cells were not altered by cryopreservation and storage at -196 degrees C up to 1 year. Cryopreserved reference lymphocytes provided a stable standard for use as stimulating cells in mixed lymphocyte cultures and could be irradiated or treated with mitomycin C prior to being frozen, without alteration of their function as stimulator cells. Cryopreserved cells thus appeared to be utilizable in the sequential measurement of the immune competence of humans.

B-Lymphocytes↗
PubMed1978

Successful engraftment of cryopreserved autologous bone marrow in patients with malignant lymphoma.

In an effort to evaluate the possible utility of cryopreserved autologous bone marrow infusions in man, 22 patients with malignant lymphoma resistant to conventional chemotherapy were treated with high-dose chemotherapy. This was followed in 12 patients by an infusion of their cryopreserved autologous bone marrow; 10 patients received chemotherapy alone and serve as controls. Following chemotherapy, severe leukopenia (less than 100 leukocytes/mm3) lasted 6-10 (median 8) days in patients receiving cryopreserved marrow, compared to 10-29 (median 16) days in controls (p less than 0.001). Recovery to 1000 leukocytes/mm3 occurred 10-18 (median 13) days after chemotherapy in autograft recipients but was delayed until 12-38 (median 23) days after chemotherapy in controls (p less than 0.001). Autografted patients also recovered granulocyte and platelet function significantly faster and had significantly fewer febrile days after chemotherapy than did controls. Cryopreserved autologous bone marrow infusions can hasten hemopoietic recovery in man after high-dose chemotherapy; this earlier reconstitution may be of clinical benefit to the patient.

Adolescent↗
PubMed2026

Mitochondrial uncoupler BAM15 attenuates cryopreservation-induced damage in human sperm by stabilizing mitochondrial homeostasis†.

Human sperm cryopreservation is essential for sperm banking and assisted reproduction, yet freeze-thaw stress promotes oxidative injury that reduces motility and damages the acrosome and nuclear DNA. Here, we tested whether the mitochondrial uncoupler BAM15 improves post-thaw human sperm quality and examined mechanisms linked to mitochondrial homeostasis. Ejaculates were cryopreserved using a standard protocol supplemented with graded concentrations of BAM15. After thawing, total and progressive motility and viability were assessed. Flow cytometry quantified the DNA fragmentation index and the proportion of high DNA stainability cells. Mitochondrial membrane potential, intracellular reactive oxygen species, and lipid peroxidation were measured to evaluate mitochondrial function and oxidative status. Ultrastructural preservation of the acrosome, plasma membrane, midpiece mitochondria, and flagellar axoneme was examined by transmission electron microscopy. Compared with untreated controls, BAM15 increased total and progressive motility and improved viability. BAM15 reduced DNA fragmentation and decreased high DNA stainability, indicating enhanced genomic integrity. Consistently, BAM15 improved mitochondrial membrane potential while suppressing intracellular reactive oxygen species and lipid peroxidation, supporting attenuation of freeze-thaw oxidative damage. Transmission electron microscopy further revealed more continuous acrosomal and plasma membranes, fewer swollen or vacuolated midpiece mitochondria, and improved preservation of axonemal architecture. Collectively, these findings identify BAM15 as a promising cryopreservation supplement that stabilizes mitochondrial homeostasis and improves the functional and structural quality of human sperm after thawing.

Humans↗
PubMedDate not supplied

Evaluation of platelet cryopreservation techniques by isolated kidney perfusion.

The isolated perfused rabbit kidney, a model system used previously to assess platelet function, was adapted for evaluation of human platelet cryopreservation techniques. A new, simple, efficient device for controlling cooling rates before, during, and after freezing was used. Platelet concentrates frozen with 5 per cent dimethyl sulfoxide (DMSO) under different conditions were the most effective of those tried in maintaining the hemostatic function and vascular integrity of perfused kidneys. Our studies indicate that the isolated, perfused rabbit kidney can be used to evaluate platelet cryopreservation techniques and is potentially adaptable for studies of organ cryopreservation.

Animals↗
PubMed1979

A modified assay for studies of cultured granulocyte precursors: cryopreservation of stimulating mononuclear cells.

Mononuclear cells (MNC), isolated from peripheral blood of healthy donors, were cryoprotected by dimethyl sulfoxide and stored in liquid nitrogen. Colony-stimulating factor (CSF), produced by cryopreserved MNC, was compared with that of nonfrozen controls in a double-layer agar system with human bone marrow as target cells. Our results indicate that cryopreserved MNC retain their ability to stimulate myelopoiesis-committed stem cells after freezing. In addition, evidence was obtained that CSF of feeder layers changes, depending on the duration of preincubation and cell concentration. In a system where either stimulating or target cells are cryopreserved the dynamics of interactions between normal or abnormal cell lines can thus be studied.

Blood Preservation↗
PubMed1977

Ultrastructure and viability of cryopreserved Plasmodium falciparum.

Cryopreserved chimpanzee erythrocytes infected with Plasmodium falciparum were examined by electron microscopy directly and after a period of culture. Light microscopic observations on the viability of cryopreserved parasites in culture were also made. Parasitaemia data from a chimpanzee infected with parasites cryostored for more than 2(1/2) years are presented. Varying degrees of damage were apparent as the parasites were examined in vitro, for viability, and by electron microscopy. Schizonts appear not to survive the process, whereas the ability of "ring" trophozoite forms to develop into large trophozoites in vitro varies from 20% to 100% in different preparations. Cryopreserved material examined by electron microscope showed degenerative changes in most of the parasites, but after 72 h of cultivation most of the parasites appeared normal. These findings suggest that some organisms may sustain reversible damage during cryostorage.

Animals↗
PubMed2026

Beyond survival: Testicular tissue cryopreservation for fertility preservation during childhood cancer, experiences from a single center.

INTRODUCTION: Approximately 30% of all male childhood cancer survivors receive gonadotoxic therapy increasing their risk for infertility. The only current option for potential fertility preservation in prepubertal males is through testicular tissue cryopreservation (TTC), available through clinical trials. Our aim was to evaluate our experience and identify barriers to enrollment in the TTC study. METHODS: A retrospective review was conducted to describe a single center experience with TTC, including an assessment of factors impacting the decision to enroll or not enroll in the TTC trial. RESULTS: A total of 61 patients were found to be eligible for enrollment in the TTC trial, with 23 patients (34%) choosing to enroll and undergo testicular tissue cryopreservation including 19 (83%) oncology patients. In patients who underwent TTC there were no unexpected delays in starting disease directed therapy. For families that declined enrollment, many did not see fertility preservation as a priority at the time of counseling. (n = 17, 44%). Other barriers included concerns about post-operative complications and a feeling of being overwhelmed at the time of counseling. DISCUSSION: Despite being the only preservation option for pre-pubertal boys facing risk of infertility, TTC enrollment was not pursued by a majority of eligible patients and families. Identifying potential barriers or misconceptions, particularly in regard to post operative complication risks, offer opportunities for improved counseling and education. Early introduction of the Fertility Navigation team allows for procedural coordination success and access for patients that are interested. Success is not measured in how many patients proceed with TTC, rather that all patients have the opportunity for counseling and discussion. CONCLUSION: TTC can be offered and completed safely without delays in disease-directed treatment timelines. A Fertility Navigation Program allows for equal opportunity for education and enrollment on TTC protocol which should be the goal for pediatric institutions, though is not pursued by many patients. A majority of those who declined the study did not see fertility as a priority at the time of treatment while others were concerned regarding peri-operative risks.

Humans↗
PubMed2018

Chromatin architecture changes and DNA replication fork collapse are critical features in cryopreserved cells that are differentially controlled by cryoprotectants.

In this work, we shed new light on the highly debated issue of chromatin fragmentation in cryopreserved cells. Moreover, for the first time, we describe replicating cell-specific DNA damage and higher-order chromatin alterations after freezing and thawing. We identified DNA structural changes associated with the freeze-thaw process and correlated them with the viability of frozen and thawed cells. We simultaneously evaluated DNA defects and the higher-order chromatin structure of frozen and thawed cells with and without cryoprotectant treatment. We found that in replicating (S phase) cells, DNA was preferentially damaged by replication fork collapse, potentially leading to DNA double strand breaks (DSBs), which represent an important source of both genome instability and defects in epigenome maintenance. This induction of DNA defects by the freeze-thaw process was not prevented by any cryoprotectant studied. Both in replicating and non-replicating cells, freezing and thawing altered the chromatin structure in a cryoprotectant-dependent manner. Interestingly, cells with condensed chromatin, which was strongly stimulated by dimethyl sulfoxide (DMSO) prior to freezing had the highest rate of survival after thawing. Our results will facilitate the design of compounds and procedures to decrease injury to cryopreserved cells.

Cell Survival↗
PubMed1976

Cryopreservation of human lymphocyte function as measured by in vitro assays.

Methods are described by which cryopreserved cells can be utilized in a number of in vitro assays. On a per cell basis, nearly total recovery of function can be demonstrated for lymphocyte transformation (mitogens, antigens and MLC) and rosette-forming cells. Excellent recovery of mononuclear cell production of leukocyte migration inhibitory factor was also observed. Highly reproducible activity was recovered on a per lymphocyte basis in lymphocyte cytotoxicity but with a definite decrement in the percentage recovery. Both for longitudinal studies of immune function and for standardization of these assays in one or more laboratories such cryopreserved cells are ofimmense value and should be widely utilized.

Blood Specimen Collection↗
PubMedDate not supplied

Cryopreservation of human lymphoid cells from various tissues.

Normal and pathological lymphoid cells, collected from different sources, were cryopreserved using a programmed freezing procedure. With this cryopreservation technique, the percentage of T and B cell surface markers and the proliferative response to mitogens were not influenced by 21 and 35 days of storage in liquid nitrogen. The recovery percentage of the lymphoid cells was satisfactory when fetal calf serum was added, as a protein source, to the medium during freezing and thawing phases, while a very low percentage of cells was recovered if the fetal calf serum was omitted.

Antigens, Surface↗
PubMed1979

In vitro induction of cytotoxic effector cells against human neoplasms. I. Sensitization conditions and effect of cryopreservation on the induction and expression of cytotoxic responses to allogeneic leukemia cells.

Peripheral blood lymphocytes (PBL) from normal human donors were sensitized in vitro against allogeneic human acute myelocytic leukemia (AML) cells by means of an unidirectional mixed lymphocyte-tumor cell culture (MLTC) technique. The cytotoxic responsiveness of the sensitized lymphocytes, as determined in vitro by the 51Cr-release assay, varied among individual lymphocyte donors and was greatly dependent on the sensitization culture conditions. Induction of cytotoxic effector cells was augmented appreciably by adding to the cultures minute amounts of the immunopotentiating agent MER-BCG. Responding lymphocytes and stimulating leukemia cells cryopreserved for several weeks in liquid nitrogen were as effective as fresh cells in generating effector lymphocytes; the cytotoxic capacity of already sensitized lymphocytes was fully retained by cryopreservation. The implications of these findings for possible clinical employment of in vitro sensitized lymphocytes in adoptive immunotherapy of cancer are discussed.

Adult↗
PubMed2026

Cryopreservable dopaminergic progenitors derived from human iPSCs with accelerated loss of pluripotency and early functional restoration in Parkinsonian rats.

Midbrain dopaminergic progenitors (mDAp) derived from human pluripotent stem cells have demonstrated promising safety and efficacy in Phase I clinical transplantation trials for Parkinson's disease (PD). To further improve the translational potential of this approach, strategies that accelerate the loss of pluripotency, increase the yield of in vitro mDA progenitors, and promote post-transplantation neurite outgrowth may be beneficial. Here, we developed an optimized protocol building upon our previously established neural induction method, with further refinements, to efficiently convert induced pluripotent stem cells (iPSCs) into mDA neurons. Additionally, we applied n-butylidenephthalide, which selectively reduced pluripotency-associated gene expression and enhanced neurite outgrowth during differentiation. With these improved techniques, 71% of iPSCs differentiated into mDA neurons, showing burst dopamine secretion and phasic electrophysiological activities with external stimuli. To evaluate the safety, cryopreserved mDAp were transplanted into immunodeficient mice. No teratoma or neural tumor was observed within 24 weeks post-implantation. In 6-OHDA PD rats, mDAp survived and differentiated into mDA neurons in the host striatum within eight weeks post-transplantation, leading to significant functional recovery. The current differentiation process, therefore, enables the generation of cryopreservable, off-the-shelf mDAp, with accelerated loss of the pluripotency marker OCT4 in vitro and early functional recovery following transplantation.

Journal Article↗
PubMedDate not supplied

Cryopreservation of human lymphocytes: a brief review and evaluation of an automated liquid nitrogen freezer.

Successful cryopreservation of human lymphocytes has been previously described. Cryopreserved lymphocytes are useful for a variety of in vitro immunologic studies. This study was performed to determine the applicability and/or advantages of using a programmable freezing system, and compares glycerol versus dimethyl sulfoxide (DMSO) at varying concentrations on post-thaw viability, E-rosetting and immunoglobulin fluorescence. Prefreeze T and B lymphocyte percentages were determined. Cells were then frozen in varying concentrations of glycerol and DMSO. Optimum cryoprotectant type and concentration was determined. Lymphocytes from seven individuals were frozen by the batch method in a mechanical freezer and with the automated liquid nitrogen injection system. Data on post-thaw T and B percentages and viability revealed 10% DMSO and liquid nitrogen control freezing method at 1 C/minute as the best conditions for lymphocyte preservation as reflected by post-thaw in vitro testing.

Blood Preservation↗
PubMed1979

The survival of cryopreserved human bone marrow stem cells.

Two methods for cryopreservation of bone marrow stem cells were compared using bone marrow obtained from 36 patients. Included in this group were 21 persons with the diagnosis of leukaemia including 14 either with acute myeloid or lymphoblastic leukaemia in remission following intensive remission induction chemotherapy. After freeze-preservation and reconstitution, all marrow samples were tested for nucleated cell (NC) recovery and grown on agar to assess colony forming units (CFUC) and cluster forming units in culture (CluFUc). A slow dilution reconstitution method using freezing media containing AB negative plasma resulted in recovery of 85% of the CFUc activity of fresh marrow. This result was significantly better than the 47% CFUc recovery obtained when freezing media without plasma and a rapid dilution reconstitution technique were used. NC recoveries following slow dilution (51%) and rapid dilution (44%) were not significantly different. CluFUc were disproportionately reduced compared with CFUc although yielding similar results with both methods (26% and 32%). No correlation was found for either method between CFUc and NC recovery or between CFUc and CluFUc recovery in cryopreserved bone marrow.

Adolescent↗