title: [Comparison of the activity and yield rate of osteoblast obtained by different digestion methods].
abstract: OBJECTIVE
  To compared the activity and yield rate of osteoblast obtained by different collagenase digestion methods, to find a better way to extract osteoblast for the experimental researches of osteoporosis.
  
  
  METHODS
  Ten 24-hour-old SD rats were were euthanized. The cranium of rats were removed and cuted into blocks of 1 mm x 1 mm size. After digested by trypsin for 15 min, all the cranium were divided into two equal parts, and randomly divided into two groups which would be digested by type I collagenase and type II collagenase separately for two times. The rat cells of the two groups were cultured in thermostat incubator with 5% CO2 under the condition of 37 degrees C. The primary culture osteoblasts were counted by using a haemacytometer after digestion and 72 hours later. The second generation osteoblasts cultured 48 h were dyed by NBT/BCIP staining solution, and were detected by quantitative measurement with PNPP.
  
  
  RESULTS
  The cells had irregular shapes. The results of cell counting showed that the cell number of type I group was larger than type 11 group. Alkaline phosphatase dyeing were positive. Detecting of alkaline phosphatase using the method of PNPP showed that the absorbance value in type I group were higher than type II group (P<0.05).
  
  
  CONCLUSIONS
  Two types of collagenase are both suitable for the in vitro culture of rat osteoblasts. The activity and yield rate of osteoblasts in type I group are higher which could provide more stable seed cells for the treatment of osteoporosis.
authors.affiliation: Shi's Center of Orthopedics and Traumatology, Shuguang Hospital Affiliated to Shanghai University of TCM, Shanghai 201203, China.
authors.forename: Ling-hui
authors.initials: LH
authors.lastname: Li
authors.forename: Dao-Fang
authors.initials: DF
authors.lastname: Ding
authors.forename: Guo-Qing
authors.initials: GQ
authors.lastname: Du
authors.forename: Hui-Hao
authors.initials: HH
authors.lastname: Wang
authors.forename: Hong-Sheng
authors.initials: HS
authors.lastname: Zhan
mesh_terms: D000818:Animals; D002452:Cell Count; D018929:Cell Culture Techniques / Q000379:methods*; D017364:Collagenases / Q000378:metabolism*; D005260:Female; D008297:Male; D010006:Osteoblasts / Q000166:cytology*; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D017364:Collagenases