Cryopreservation of human mucosal leukocytes
Cryopreservation of human mucosal leukocytes
Abstract BackgroundUnderstanding how leukocytes in the cervicovaginal and colorectal mucosae respond to pathogens, and how medical interventions affect these responses, is important for developing better tools to prevent HIV and other sexually transmitted infections. An effective cryopreservation protocol for these cells following their isolation will make studying them more feasible. Methods and findingsTo find an optimal cryopreservation protocol for mucosal mononuclear leukocytes, we compared cryopreservation media and procedures using human vaginal leukocytes and confirmed our results with endocervical and colorectal leukocytes. Specifically, we measured the recovery of viable vaginal T cells and macrophages after cryopreservation with different cryopreservation media and handling procedures. We found several cryopreservation media that led to recoveries above 75%. Limiting the number and volume of washes increased the fraction of cells recovered by 10-15%, possibly due to the small cell numbers in mucosal samples. We confirmed that our cryopreservation protocol also works well for both endocervical and colorectal leukocytes.Cryopreserved leukocytes had slightly increased cytokine responses to antigenic stimulation relative to the same cells tested fresh. Additionally, we tested whether it is better to cryopreserve endocervical cells on the cytobrush or in suspension. ConclusionsLeukocytes from cervicovaginal and colorectal tissues can be cryopreserved with good recovery of functional, viable cells using several different cryopreservation media. The number and volume of washes has an experimentally meaningful effect on the percentage of cells recovered. We provide a detailed, step-by-step protocol with best practices for cryopreservation of mucosal leukocytes.
Shu Zhiquan、Ferre April L.、Fialkow Michael、Kirby Anna C.、Adams Waldorf Kristina M.、McElrath M. Juliana、Hladik Florian、Levy Claire N.、Baker Chris A. R.、Veazey Ronald S、Gao Dayong、Briesen Hagen von、Hughes Sean M.、Sinclair Elizabeth、Hartig Heather、Shacklett Barbara L、Lentz Gretchen、Germann Anja、Fang Cifeng、Dezzutti Charlene S.
Department of Mechanical Engineering, University of Washington||School of Mechanical and Materials Engineering, Washington State University, EverettDepartment of Medical Microbiology and Immunology, School of Medicine, University of California at DavisDepartment of Obstetrics and Gynecology, University of WashingtonDepartment of Obstetrics and Gynecology, University of WashingtonDepartment of Obstetrics and Gynecology, University of WashingtonVaccine and Infectious Disease Division||Department of Medicine, University of Washington||Department of Laboratory Medicine, University of Washington||Department of Global Health, University of WashingtonDepartment of Obstetrics and Gynecology, University of Washington||Vaccine and Infectious Disease Division||Department of Medicine, University of WashingtonDepartment of Obstetrics and Gynecology, University of WashingtonDivision of Experimental Medicine, Department of Medicine, San Francisco General Hospital, University of California San Francisco||Core Immunology Lab, University of California San FranciscoDivision of Comparative Pathology, Tulane National Primate Research Center, Tulane University School of MedicineDepartment of Mechanical Engineering, University of Washington||Department of Bioengineering, University of WashingtonDivision of Medical Biotechnology, Fraunhofer Institute for Biomedical Engineering IBMTDepartment of Obstetrics and Gynecology, University of WashingtonDivision of Experimental Medicine, Department of Medicine, San Francisco General Hospital, University of California San FranciscoDepartment of Medicine, University of California San FranciscoDepartment of Medical Microbiology and Immunology, School of Medicine, University of California at Davis||Department of Medicine, Division of Infectious Diseases, School of Medicine, University of California at DavisDepartment of Obstetrics and Gynecology, University of WashingtonDivision of Medical Biotechnology, Fraunhofer Institute for Biomedical Engineering IBMTDepartment of Mechanical Engineering, University of WashingtonSchool of Medicine, University of Pittsburgh||Magee-Womens Research Institute
医学研究方法预防医学基础医学
Shu Zhiquan,Ferre April L.,Fialkow Michael,Kirby Anna C.,Adams Waldorf Kristina M.,McElrath M. Juliana,Hladik Florian,Levy Claire N.,Baker Chris A. R.,Veazey Ronald S,Gao Dayong,Briesen Hagen von,Hughes Sean M.,Sinclair Elizabeth,Hartig Heather,Shacklett Barbara L,Lentz Gretchen,Germann Anja,Fang Cifeng,Dezzutti Charlene S..Cryopreservation of human mucosal leukocytes[EB/OL].(2025-03-28)[2025-05-05].https://www.biorxiv.org/content/10.1101/039578.点此复制
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