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Assessment of Protein Profiles of RNAlater Stored and Fresh PBMC Cells Using Different Protein Extraction Buffers

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Abstract

For proteome analyses, the tissue samples are mostly preserved either snap frozen or formalin-fixed, paraffin-embedded form. Use of RNAlater—a non-toxic solution primarily used to stabilize the RNA content of samples—in tissue preservation for proteome analysis recently described equally reliable with snap-frozen preservation in human tissues. Even though RNALater storage has great potential in the preservation of Peripheral Blood Mononuclear Cells (PBMC), its impact on the results of proteome analysis is poorly described at qualitative and quantitative measures. The present study investigated protein profiles of RNAlater preserved and fresh PBMCs using three extraction buffers viz. Triton X-100, RIPA and SDS. Proteins are separated in SDS-PAGE and quantified using densitometry. On an average 19.3 bands from fresh and 15.6 bands from RNAlater storage cells were obtained with a molecular weight ranging from 25 to > 250 kDa. RNAlater storage generated a fewer number and lesser quantity of low molecular weight proteins while yielded a similar or high quantity of high molecular weight protein fractions. The principal component analysis showed that Triton X-100 is inferior as compared to SDS and RIPA with respect to their protein bands and quantity yielded. While RNAlater is effective in preserving PBMC for proteome analysis, our findings warrant caution in its use in proteomics experiments especially if the target is low molecular weight proteins.

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Funding

This research work was funded by DST-SERB, Government of India; Project Number DST_SERB_ECR/2017/000761.

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Authors and Affiliations

  1. ICAR-Central Island Agricultural Research Institute, Port Blair, Andaman & Nicobar Islands, India

    R. R. Alyethodi, S. Karthik, K. Muniswamy, S. K. Ravi, P. Perumal, D. Bhattacharya, P. A. Bala, A. K. De, T. Sujatha, Jai Sunder & A. Kundu

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  1. R. R. Alyethodi
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  2. S. Karthik
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  3. K. Muniswamy
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  4. S. K. Ravi
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  6. D. Bhattacharya
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Correspondence to R. R. Alyethodi.

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Research has been approved by the Institute Animal Ethics Committee (IAEC) of the ICAR-Central Institute for Research on Cattle, Meerut, UP, India under CPCSEA guidelines. This article does not contain any studies with human participants performed by any of the authors.

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Alyethodi, R.R., Karthik, S., Muniswamy, K. et al. Assessment of Protein Profiles of RNAlater Stored and Fresh PBMC Cells Using Different Protein Extraction Buffers. Protein J 39, 291–300 (2020). https://doi.org/10.1007/s10930-020-09888-y

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