Cryostor cs5

Manufactured by BioLife Solutions

Sourced in United States

CryoStor-CS5 is a serum-free, protein-free, and chemically-defined cryopreservation medium designed for the preservation of cells and tissues at cryogenic temperatures. It is formulated to provide optimal cell recovery and viability following cryogenic storage.

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Lab products found in correlation

Clinimacs device, by Miltenyi Biotec (2 mentions) Rhil 15, by CellGenix (2 mentions) Knockout sr, by Thermo Fisher Scientific (2 mentions) Rhil 7, by CellGenix (2 mentions) Rhil 21, by Miltenyi Biotec (2 mentions) Cts dynabeads cd3 cd28, by Thermo Fisher Scientific (2 mentions) Sepax 2rm device, by Cytiva (2 mentions) X vivo 15, by Lonza (2 mentions) Anti nkp46, by R&D Systems (1 mentions) Recombinant human il 18, by R&D Systems (1 mentions) Recombinant human il 2, by Novartis (1 mentions) Deoxyribonuclease 1 from bovine pancreas, by Merck Group (1 mentions) Collagenase from clostridium histolyticum, by Merck Group (1 mentions) Ack lysis buffer, by Lonza (1 mentions) Gentlemacs dissociator system, by Miltenyi Biotec (1 mentions) Spectra optia, by Terumo BCT (1 mentions) Elutra, by Terumo BCT (1 mentions) Heat inactivated fetal bovine serum, by Merck Group (1 mentions) Dulbecco s phosphate buffered saline, by Corning (1 mentions) Ficoll paque plus, by GE Healthcare (1 mentions)

6 protocols using cryostor cs5

Anti-B7-H3 CAR-T Cell Production

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CD4+ and CD8+ T cells were isolated from patient apheresis products using the CliniMACS device (Miltenyi Biotec). A 1:1 mixture of CD4-enriched and CD8-enriched cell fractions were then pooled, suspended in X-Vivo 15 (Lonza) media supplemented with 2% KnockOut SR (Life Technologies), 5 ng/mL rhIL-7 (CellGenix), 0.5 ng/mL rhIL-15 (CellGenix) and 10 ng/mL rhIL-21 (Miltenyi Biotec), initiated into culture in a G-Rex 100MCS vessel (Wilson-Wolf), and stimulated using CD3/CD28 CTS® Dynabeads (Life Technologies). Cell products were then transduced with a GMP-grade SIN (self-inactivating) lentivirus encoding the anti-B7-H3CAR, the methotrexate-resistant human DHFR mutein huDHFRdm, and the cell-surface marker EGFRt. On day 3 of culture, 50nM methotrexate (Mylan) was added to the culture vessel to select for cells possessing DHFRdm. On day 7 of culture, CD3/CD28 CTS® beads were removed from the cell suspension using the Dynamag CTS device. Following 10–13 days in culture, patient cells were harvested and washed using the Sepax 2RM device (Cytiva) and resuspended in CryoStor-CS5 (Biolife Solutions) for cryopreservation in CellSeal closed-system vials (Sexton).

Vitanza N.A., Wilson A.L., Huang W., Seidel K., Brown C., Gustafson J.A., Yokoyama J.K., Johnson A.J., Baxter B.A., Koning R.W., Reid A.N., Meechan M., Biery M.C., Myers C., Rawlings-Rhea S.D., Albert C.M., Browd S.R., Hauptman J.S., Lee A., Ojemann J.G., Berens M.E., Dun M.D., Foster J.B., Crotty E.E., Leary S.E., Cole B.L., Perez F.A., Wright J.N., Orentas R.J., Chour T., Newell E.W., Whiteaker J.R., Zhao L., Paulovich A.G., Pinto N., Gust J., Gardner R.A., Jensen M.C, & Park J.R. (2022). Intraventricular B7-H3 CAR T Cells for Diffuse Intrinsic Pontine Glioma: Preliminary First-in-Human Bioactivity and Safety. Cancer Discovery, 13(1), 114-131.

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Expansion of Purified Natural Killer Cells

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The expansion of isolated pNK cell was performed with a GMP-compliant method at CHA Biotech (Seongnam, Korea). The cells were seeded on a γ-globulin (Greencross, Yongin, Korea) and anti-NKp46 (R&D Systems, Minneapolis, MN, USA)-coated flask and cultured in Alys505NK serum-free medium (CSTI, Sendai, Japan) supplemented with 1000 IU/mL recombinant human IL-2 (Novartis, Basel, Switzerland), 50 ng/mL recombinant human IL-18 (R&D system, Minneapolis, MN, USA), and 5% heat-inactivated autologous plasma. Fresh culture medium was added every 1 to 3 days depending on the cell density (2 × 10⁶ cells/mL). On Day 6, the cells were transferred to a culture bag (NIPRO, Osaka, Japan), and cultured for 14 days. The pNK cells were cryopreserved with Cryostor CS5 (BioLife Solutions, Bothell, WA, USA).

Jung D., Baek Y.S., Lee I.J., Kim K.Y., Jang H., Hwang S., Jung J., Moon Y.W., Park K.S., Choi Y.S, & An H.J. (2021). Ex vivo expanded allogeneic natural killer cells have potent cytolytic activity against cancer cells through different receptor-ligand interactions. Journal of Experimental & Clinical Cancer Research : CR, 40, 333.

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Engineered Antigen-Specific T Cell Therapy

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CD4⁺ and CD8⁺ T cells were isolated from patient apheresis products using the CliniMACS device (Miltenyi Biotec). A 1:1 mixture of CD4-enriched and CD8-enriched cell fractions were then pooled, suspended in X-Vivo 15 (Lonza) media supplemented with 2% KnockOut SR (Life Technologies), 5 ng/mL rhIL7 (CellGenix), 0.5 ng/mL rhIL15 (CellGenix), and 10 ng/mL rhIL-21 (Miltenyi Biotec), initiated into culture in a G-Rex 100MCS vessel (Wilson-Wolf), and stimulated using CD3/CD28 CTS Dynabeads (Life Technologies). Cell products were then transduced with a GMP-grade SIN (self-inactivating) lentivirus encoding the anti-B7-H3 CAR, the MTX-resistant human DHFR mutein huDHFRdm, and the cell-surface marker EGFRt. On day 3 of culture, 50 nmol/L MTX (Mylan) was added to the culture vessel to select for cells possessing DHFRdm. On day 7 of culture, CD3/CD28 CTS beads were removed from the cell suspension using the Dynamag CTS device. Following 10 to 13 days in culture, patient cells were harvested and washed using the Sepax 2RM device (Cytiva) and resuspended in CryoStor-CS5 (Biolife Solutions) for cryopreservation in CellSeal closed-system vials (Sexton).

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Isolation and Cryopreservation of Decidua Basalis and PBMC

Cited 2 times

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Freshly collected decidua basalis was washed with cold PBS and processed using methods previously described (37 (link)). Briefly, tissue was minced and dissociated in RPMI containing 1 mg/mL of Collagenase from Clostridium histolyticum (Sigma Aldrich, St. Louis, MO) and 1 μg/mL Deoxyribonuclease I from bovine pancreas (Sigma Aldrich, St. Louis, MO), using the GentleMACS Dissociator system (Miltenyi Biotec Inc., San Diego, CA, USA). Homogenates were then filtered through a 100-μm filter; red blood cells were lysed with ACK Lysis Buffer (Lonza, Walkersville, MD), and mononuclear cells (MCs) were recovered and frozen in CryoStor CS5 (BioLife Solutions, Bothell, WA) and stored in liquid nitrogen before evaluation. PBMCs were isolated from peripheral blood collected in EDTA tubes using previously described standard methods (38 (link)), frozen in CryoStor CS5, and stored in liquid nitrogen prior to evaluation. PBMC samples taken on the day of pregnancy termination were used for most cases. If such a sample was not available, the closest available PBMC sample, prior to surgical pregnancy termination, was used. Full details on the PBMC samples used can be found in Supplementary Table 1.

Koenig M.R., Vazquez J., Leyva Jaimes F.B., Mitzey A.M., Stanic A.K, & Golos T.G. (2024). Decidual leukocytes respond to African lineage Zika virus infection with mild anti-inflammatory changes during acute infection in rhesus macaques. Frontiers in Immunology, 15, 1363169.

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Isolation and Purification of Immune Cell Subsets

Cited 1 time

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Peripheral blood lymphocytes (PBLs) from HLA-A2 positive donors were collected as previously described^{21 (link)}. Briefly, leukapheresis (Spectra Optia®, TerumoBCT) was performed on consenting healthy donors (Ethical license B.U.N. 143201420996) in accordance with the relevant guidelines and regulations. The leukapheresis product was processed via an elutriation procedure to collect monocytes (Elutra®, TerumoBCT). The remaining PBLs were counted and cryopreserved in 5% DMSO-containing cryopreservation medium (CryoStor® CS5, Biolife Solutions) till further use.
To study the contribution of NK cells to the observed effects, PBLs were depleted from CD19⁺ cells using anti-CD19 magnetic beads to enhance purity of NK, CD4⁺ and CD8⁺ T cells within the PBLs. Subsequently, CD4⁺ and CD8⁺ T cells were depleted with respectively anti-CD4 and anti-CD8 magnetic beads. To study the contribution of T cells to the observed effects, PBLs were selected for CD4⁺ and CD8⁺ T cells using anti-CD4 and anti-CD8 magnetic beads. Depletions and selections were performed according to the manufacturer’s instructions.

Awad R.M., De Vlaeminck Y., Meeus F., Ertveldt T., Zeven K., Ceuppens H., Goyvaerts C., Verdonck M., Salguero G., Raes G., Devoogdt N, & Breckpot K. (2023). In vitro modelling of local gene therapy with IL-15/IL-15Rα and a PD-L1 antagonist in melanoma reveals an interplay between NK cells and CD4+ T cells. Scientific Reports, 13, 18995.

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Isolation of Human CD14+ Monocytes

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Human white blood cells were collected from leukoreduction filters (Sepacell RS-2000; Fenwal Inc.) after processing of healthy volunteer donor whole blood (American Red Cross) after rinsing with 50 mL of selection buffer (2 mmol/L ethylenediaminetetraacetic acid (EDTA; J.T. Baker Inc.), 2% heat-inactivated fetal bovine serum (Sigma-Aldrich) in Dulbecco’s phosphate- buffered saline (Corning Cellgro)). The filtrate was layered onto Ficoll-Paque PLUS (GE Healthcare) to isolate mononuclear cells by density gradient centrifugation. Mononuclear cells treated with CD14-positive selection beads (BD Biosciences) in selection buffer and subsequent EasySep magnetic purification procedure (Stemcell Technologies) resulted in purified CD14+ monocytes (mean purity 95%), which were cryopreserved in Cryostor CS5 (Biolife Solutions). These primary human monocytes were labelled with BCECF-AM and used in adhesion assays as described above for THP-1 cells.

Rajendran N.K., Liu W., Chu C.C., Cahill P.A, & Redmond E.M. (2021). Moderate dose alcohol protects against serum amyloid protein A1-induced endothelial dysfunction via both notch-dependent and -independent pathways. Alcoholism, clinical and experimental research.

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