Comparison of different protocols for protein extraction from murine ovarian tissue: A preliminary study

Nitika Sharma, Ishwerpreet Kaur Jawanda, Thomson Soni and Vijay Prabha *

Department of Microbiology, Panjab University, Chandigarh-160014, India.
 
Research Article
International Journal of Science and Research Archive, 2023, 08(02), 172–179.
Article DOI: 10.30574/ijsra.2023.8.2.0238
Publication history: 
Received on 10 February 2023; revised on 19 March 2023; accepted on 21 March 2023
 
Abstract: 
Objective: Extraction of proteins from tissues is a key step to study the protein structure and function. There is no set of universal protocols because of the enormous chemical and physical variety of both proteins and the sample sources. Hence, in this study a comparison of different methods for protein extraction from mouse ovarian tissue was carried out.
Methods: Intravaginal dose of 20µl of Phosphate buffer saline was given to 7 mice for ten consecutive days followed by extraction of ovaries. For comparison, seven different protein extraction protocols from Protocol 1 to Protocol 7 were used. The supernatant was first quantified with Bradford assay to determine the total protein yield followed by SDS-PAGE analysis.
Results: Highest protein yield of 1.13 mg/ml was attained after supplementing 1% v/v protease inhibitor cocktail in PBS. In total, 13 protein bands were detected by the extraction protocol 2 and 11 protein bands by the extraction protocol 1. 11 protein bands were detected in the range 180 kDa to 22 kDa following Protocol 3 and 4. Extraction with lysis buffer 1 resulted in 7 visible protein bands of molecular weight 182 kDa to 35 kDa range. Only 6 protein bands were observed in the range 182 kDa - 35 kDa following lysis buffer 2 method. For lysis buffer 3, ten visible protein bands were detected in range of 195 kDa - 13 kDa.
Conclusion: Based on the protein extraction efficiency of Protocol 1, optimal combination of Phosphate buffer saline (pH 7.2) and protease inhibitor cocktail followed by hand-homogenization is suggested for protein extraction from mouse tissues.
 
Keywords: 
Protein extraction; Ovarian tissue; Homogenization; Lysis buffer
 
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