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Tissue processing

TN Thanh Loc Nguyen
YC Youngjin Choi
JI Jihye Im
HS Hyunsu Shin
NP Ngoc Man Phan
MK Min Kyung Kim
SC Seung Woo Choi
JK Jaeyun Kim
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Spleens were excised and processed by mechanical disruption using a 70 µm cell strainer. Cells were then centrifuged for 5 min, 400 × g, 4 °C and treated with ammonium-chloride-potassium (ACK) lysing buffer (Lonza) for 4 min to remove red blood cells. Splenocytes were then filtered through a 40 µm cell strainer and washed in cold PBS. Spinal cords were dissociated in PBS containing 1 mg/mL collagenase type IV and 20% EDTA/trypsin and incubated at 37 °C for 20 min. RPMI 1640 containing 10% fetal bovine serum was added to each sample to inhibit enzymatic activity, and the cells were filtered through a 40 µm cell strainer. The cells were collected by centrifugation according to standard protocol.

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