Application of retrograde dissection method for isolation of bone marrow cells from rat femurs and tibiae

作者：Li, C. M.; Fu, B. M.^*; Zhang, L. C.; Tang, B.; Zhu, L.; Zhao, Y.; Zhang, J.^*

来源：Genetics and Molecular Research, 2016, 15(2): 15028178.

DOI：10.4238/gmr.15028178

• Retrograde dissection
• Tibiae
• Femurs
• Rat
• Bone marrow cells

Currently, there is no practical and efficient method for the isolation of bone marrow cells (BMCs) from rat femurs and tibiae. Here, we attempted to develop a rapid, simple, effective, and non-contaminating method for the isolation of BMCs from rat femurs and tibiae. Rat femurs and tibiae were dissected from the ankle to the hip joint; subsequently, a three-step "locate-slide-twist" procedure was performed using scissors and forceps to remove the femurs and tibiae completely, from the surrounding musculature. The bones were flushed with phosphate-buffered saline to harvest BMCs. The femurs and tibiae were dissected in 1.8 +/- 0.6 min, and the BMC suspension preparation time was 13.1 +/- 2.3 min. The bone marrow cavities did not incur any fractures or injuries during the isolation. Culture of harvested BMCs for 72 h led to a significant increase in cell number from 4.4 +/- 0.3 x 10(6) to 6.9 +/- 0.7 x 10(6) (P < 0.01) with no significant decrease in viability (98.1 +/- 0.6% nu s 96.2 +/- 1.1%; P > 0.05). Microscopic examination of the isolated BMCs after the 72-h incubation period revealed the no-microbial or muscle cell contamination. Furthermore, flow cytometry revealed that cultured BMCs (72-h culture) grew well. Here, we have reported a rapid, simple, effective, and non-contaminating method for the isolation of BMCs from rat femurs and tibiae by using retrograde dissection. This method can be used to harvest a large number of viable BMCs without the risk of contamination from muscle and connective tissues.

• 出版日期2016
• 单位昆明医科大学