Cryopreserved samples were processed as previously described [78 (link)]. Briefly, intestinal tissue samples were quickly thawed and washed in T-cell media that consists of: RPMI medium plus 10% FBS (Corning), 1X GlutaMax, 10 mM HEPES, 1X MEM NEAA, 1 mM sodium pyruvate (Gibco), 100 I.U/mL penicillin, and 100 μg/mL streptomycin. Next, intestinal tissue was incubated overnight in the same media with 1 μg/mL DNase and 100 μg/mL collagenase A. Tissue dissociation was performed on the gentleMACS Octo Dissociator with heaters (Miltenyi Biotec) using the heated human tumor protocol 1. Tissue was then filtered through a 70-μm nylon mesh cell strainer (Sigma) to make a single-cell suspension.
70 μm nylon mesh cell strainer
Manufactured by Merck Group
The 70-μm nylon mesh cell strainer is a laboratory filtration tool designed to isolate and separate cells from a heterogeneous solution. It features a nylon mesh with 70-micrometer pores that allow the passage of single cells while retaining larger cell aggregates, debris, and clumps.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Corning (2 mentions)
Gentlemacs octo dissociator, by Miltenyi Biotec (2 mentions)
Sodium pyruvate, by Thermo Fisher Scientific (1 mentions)
Mem neaa, by Thermo Fisher Scientific (1 mentions)
Glutamax, by Thermo Fisher Scientific (1 mentions)
Ca2 and mg2, by Merck Group (1 mentions)
2 protocols using 70 μm nylon mesh cell strainer
1
Intestinal Tissue Dissociation and Single-Cell Suspension
2
Intestinal Tissue Dissociation Protocol
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Intestinal tissue samples were quickly thawed and washed in RPMI medium plus 10% FBS (Corning), 1× GlutaMax, 10 mM Hepes, 1× MEM nonessential amino acids, 1 mM sodium pyruvate, 100 IU/ml penicillin, and 100 μg/ml streptomycin. Next, intestinal tissue was incubated overnight in the same media with 1 μg/ml DNase 1 and 100 μg/ml collagenase A. Tissue dissociation was performed on the gentleMACS Octo Dissociator with heaters (Miltenyi Biotec) using the heated human tumor protocol 1. Tissue was then filtered through a 70-μm nylon mesh cell strainer (Sigma). A single-cell suspension was made by washing in DPBS without Ca2+ and Mg2+ (Sigma).
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