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Naser Kalhor ¹, Hoda Fazaeli ¹, Reza Tabatabaei Qomi ¹, Mohsen Sheykh Hasan ¹, Mahdieh Ghiasi ¹

Affiliations
1 Jihad Daneshgahi Infertility Center, shabnam avenue, isar square, Qom, IR

Abstract

Problems: Contamination of cell lines and biological products is one of the major problems of cell culture techniques. Rapid detection of mycoplasma contamination in cell culture is an important part of controlled laboratory. In order to existence of infected cells, leading to unreliable results, the need for a reliable method for laboratories is unavoidable. Polymerase chain reaction (PCR), a technique of rapid, sensitive and specific detection of bacteria is considered as a precise method to detect contamination. The aim of this study was to evaluate the efficacy of PCR in the detection of contaminants in cell cultures and other biological products.

Experimental approach: In this study, PCR techniques were optimized by use of MGSO and GPO-1 primers and16SrRNA target gene. Also the utilized PCR method was evaluated in terms of sensitivity and specificity. Finally, a simple DNA extraction and PCR of 164 cell culture of adipose tissue derived mesenchymal stem cells were done.

Findings: A 715 bp product was amplified by the primers and was confirmed by sequencing. The Reviews feature, with none of the tested DNA was amplified products. This method has a sensitivity limit of 10 copies of the target DNA. No cross-reactivity with genomic DNA of other microorganisms was observed.

Conclusion: The results of this study indicate that molecular methods based on polymerase chain reaction (PCR) have great importance due to their innate features. The observed results in this study are based on accuracy, speed, sensitivity and high specification of PCR technique according to conserved and common sequence of 16srRNA to detect Mycoplasma contamination in cell cultures.

Keywords

Mycoplasma, PCR, Contamination, Molecular Detection, Cell Culture.

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Mahdieh Ghiasi ¹, Reza Tabatabaei Qomi ¹, Naser Kalhor ¹, Hoda Fazaeli ¹, Mohammad Mehdizadeh ², Mohsen Sheykh Hasan ¹

Affiliations
1 Jihad Daneshgahi Infertility Center, Stem cell laboratory, shabnam avenue, isar square, Qom, IR
2 Dept. of Oral and Maxillofacial Surgery, Babol University of Medical Sciences, Babol, IR

Abstract

Problems: Tissue engineering (TE) is one of the modern strategies to provide a functional biological tissue equivalent to restore or improve the function of human tissues that lost by disease, traumatic events, or congenital abnormalities. Some essential factors in TE are tissue type and appropriate scaffold. suitable influence on cell viability and proliferation is One of the most important characteristics of a appropriate scaffold for tissue engineering. The purpose of this research was to comparative evaluation of adipose derived stem cells (ADSCs) proliferation rate and their viability on the five different scaffolds. Experimental approach: six different scaffolds were prepared including Alginate, Poly Lactic Glycolic Acid (PLGA), Fibrin glue (FG), Inactive Platelet-Rich Plasma (IPRP), Active Platelet-Rich Plasma (APRP) and Hydroxyapatite/β-tricalcium phosphate (HA/β-TCP). Human adipose tissue was applied as mesenchymal stem cells source. These cells were seeded in the targeted scaffolds. After 48 hours, the proliferation and viability of mesenchymal stem cells were assessed by MTT assay. The obtained results were analyzed statistically by SPSS software (p<0.05). Findings: Significant differences between cell proliferation and viability of different scaffolds have been shown. Alginate and active PRP were shown to be the most and least suitable scaffolds in terms of enhancing cell proliferation and maintaining cell viability respectively (p60;0.05). Conclusion: Our study indicated that active PRP could be the best scaffold for support of Adipose-derived stem cells proliferation in comparison with others.

Keywords

Adipose-Derived Mesenchymal Stem Cells, MTT Assay, Alginate, PRP, PLGA, Fibrin Glue, Hydroxyapatite/β-Tricalcium Phosphate (HA/β-TCP).

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