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Hepes ph 7

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HEPES pH 7.2 is a buffering agent used in cell culture and biochemical applications. It maintains a stable pH of 7.2 in aqueous solutions.

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Lab products found in correlation

Dnase 1, by AppliChem (4 mentions) Collagenase p, by Roche (2 mentions) Macs microbead, by Miltenyi Biotec (2 mentions) Gentlemacs based mechanical disruption, by Miltenyi Biotec (2 mentions) Collagenase type p, by Merck Group (2 mentions) Macs anti cd45 and anti ter119 microbeads, by Miltenyi Biotec (2 mentions) Collagenase type 2, by Merck Group (2 mentions)

4 protocols using hepes ph 7

1

Stromal Cell Enrichment from Lymph Nodes

Cited 1 time
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For stromal cell preparation, lymph nodes were torn into small pieces and collected in RPMI 1640 medium containing 2% FCS, 20 mM HEPES pH 7.2 (Lonza), 0.375 mg/ml Collagenase P (Roche) and 25 μg/ml DNaseI (Applichem). Dissociated tissue was incubated at 37 °C for 60 min, with resuspension and collection of the supernatant every 15 min. Following enzymatic digestion, cell suspensions were filtered and washed with PBS containing 0.5% FCS and 10 mM EDTA. For cell-sorting experiments, stromal cells were enriched by depleting CD45+ hematopoietic cells and TER119+ erythrocytes using MACS microbeads (Miltenyi) as described previously7 (link). Cell suspensions were directly used for staining with antibodies. For isolation of hematopoietic cells, lymph nodes were gently smashed across a 26-gauge wire mesh and washed with PBS. Cell suspensions were directly used for staining with antibodies or for ELISPOT assays.
Pikor N.B., Mörbe U., Lütge M., Gil-Cruz C., Perez-Shibayama C., Novkovic M., Cheng H.W., Nombela-Arrieta C., Nagasawa T., Linterman M.A., Onder L, & Ludewig B. (2020). Remodeling of light and dark zone follicular dendritic cells governs germinal center responses. Nature immunology, 21(6), 649-659.
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2

Stromal Cell Enrichment from Lymph Nodes

Cited 1 time
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For stromal cell preparation, lymph nodes were torn into small pieces and collected in RPMI 1640 medium containing 2% FCS, 20 mM HEPES pH 7.2 (Lonza), 0.375 mg/ml Collagenase P (Roche) and 25 μg/ml DNaseI (Applichem). Dissociated tissue was incubated at 37 °C for 60 min, with resuspension and collection of the supernatant every 15 min. Following enzymatic digestion, cell suspensions were filtered and washed with PBS containing 0.5% FCS and 10 mM EDTA. For cell-sorting experiments, stromal cells were enriched by depleting CD45+ hematopoietic cells and TER119+ erythrocytes using MACS microbeads (Miltenyi) as described previously7 (link). Cell suspensions were directly used for staining with antibodies. For isolation of hematopoietic cells, lymph nodes were gently smashed across a 26-gauge wire mesh and washed with PBS. Cell suspensions were directly used for staining with antibodies or for ELISPOT assays.
Pikor N.B., Mörbe U., Lütge M., Gil-Cruz C., Perez-Shibayama C., Novkovic M., Cheng H.W., Nombela-Arrieta C., Nagasawa T., Linterman M.A., Onder L, & Ludewig B. (2020). Remodeling of light and dark zone follicular dendritic cells governs germinal center responses. Nature immunology, 21(6), 649-659.
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3

Lung Lymphocyte and Stromal Cell Isolation

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Mice were sacrificed at the indicated time points and immediately perfused with PBS. Lung-infiltrating lymphocytes were isolated using mechanical disruption of the organ. For isolation of myeloid and stromal cells from the lung, the tissue was cut into small pieces, transferred into a 24-well dish filled with RPMI 1640 medium containing 2% FCS, 20 mM Hepes pH 7.2 (all from Lonza), 1 mg/ml Collagenase Type P (Sigma-Aldrich), 25 μg/ml DNaseI (AppliChem) and 1 mg/ml Collagenase Type II (Sigma-Aldrich) in combination with gentleMACS-based mechanical disruption (Miltenyi Biotech). After 30 minutes incubation at 37°C, cell suspensions were washed with PBS containing 0.5% FCS and 10 mmol/L EDTA. Stromal cell fraction enrichment was achieved by depleting hematopoietic and erythroid cells using MACS anti-CD45 and anti-Ter119 microbeads (Miltenyi Biotec).
Cupovic J., Ring S.S., Onder L., Colston J.M., Lütge M., Cheng H.W., De Martin A., Provine N.M., Flatz L., Oxenius A., Scandella E., Krebs P., Engeler D., Klenerman P, & Ludewig B. (2021). Adenovirus vector vaccination reprograms pulmonary fibroblastic niches to support protective inflating memory CD8+ T cells. Nature immunology, 22(8), 1042-1051.
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4

Lung Lymphocyte and Stromal Cell Isolation

Check if the same lab product or an alternative is used in the 5 most similar protocols
Mice were sacrificed at the indicated time points and immediately perfused with PBS. Lung-infiltrating lymphocytes were isolated using mechanical disruption of the organ. For isolation of myeloid and stromal cells from the lung, the tissue was cut into small pieces, transferred into a 24-well dish filled with RPMI 1640 medium containing 2% FCS, 20 mM Hepes pH 7.2 (all from Lonza), 1 mg/ml Collagenase Type P (Sigma-Aldrich), 25 μg/ml DNaseI (AppliChem) and 1 mg/ml Collagenase Type II (Sigma-Aldrich) in combination with gentleMACS-based mechanical disruption (Miltenyi Biotech). After 30 minutes incubation at 37°C, cell suspensions were washed with PBS containing 0.5% FCS and 10 mmol/L EDTA. Stromal cell fraction enrichment was achieved by depleting hematopoietic and erythroid cells using MACS anti-CD45 and anti-Ter119 microbeads (Miltenyi Biotec).
Cupovic J., Ring S.S., Onder L., Colston J.M., Lütge M., Cheng H.W., De Martin A., Provine N.M., Flatz L., Oxenius A., Scandella E., Krebs P., Engeler D., Klenerman P, & Ludewig B. (2021). Adenovirus vector vaccination reprograms pulmonary fibroblastic niches to support protective inflating memory CD8+ T cells. Nature immunology, 22(8), 1042-1051.
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