Chemical-Free Extraction of Functional Mitochondria Using a Microfluidic Device
| Autor: | Sung-Tzu Chen, Yu-Han Hsiao, Gou-Jen Wang, Jui-Chih Chang, Ching-Wen Li, Chin-San Liu |
|---|---|
| Rok vydání: | 2018 |
| Předmět: |
mitochondria extraction
0301 basic medicine Lysis lcsh:Engineering machinery tools and implements chip-based microfluidic device Mitochondrion Flow cytometry 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine mitochondrial function medicine lcsh:Technological innovations. Automation Membrane potential lcsh:HD45-45.2 medicine.diagnostic_test Chemistry Acridine orange General Engineering Chemical modification Staining 030104 developmental biology Membrane protein 030220 oncology & carcinogenesis Biophysics lcsh:TA213-215 |
| Zdroj: | Inventions Volume 3 Issue 4 Inventions, Vol 3, Iss 4, p 68 (2018) |
| ISSN: | 2411-5134 |
| DOI: | 10.3390/inventions3040068 |
| Popis: | This paper proposes the use of a chip-based microfluidic device to extract functional and chemical free mitochondria. A simple microfluidic device was designed and fabricated. An osteosarcoma cybrid cell line was employed to demonstrate the efficiency of the proposed microfluidic device. The membrane proteins (mitochondrial complex I-V and Tom20) and morphology of the extracted mitochondria were examined by Western blot and transmission electron microscopy (TEM), respectively. The purity and mitochondrial membrane potential of the extracted mitochondria were individually measured by 10-N-alkyl acridine orange and tetramethylrhodamine ethyl ester staining via flow cytometry. Experimental results revealed that expressed pattern of complex I&ndash V in device-extracted mitochondria was close to that of mitochondria in total cell lysis and device extraction significantly prevented chemical modification of complex IV protein via a conventional kit, although device extract similar amounts of mitochondria to the conventional kit revealed by Tom20 expression. Furthermore, purity of device-extracted mitochondria was above 93.7% and mitochondria still retained normal activity after device extraction proven by expression of mitochondrial membrane potential as well as the entire mitochondrial morphology. These results confirmed that the proposed microfluidic device could obtain functional mitochondria without structural damage. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|