Authentication of Primary Murine Cell Lines by a Microfluidics-Based Lab-On-Chip System
Abstract
1. Introduction
2. Materials and Methods
2.1. Animal Experiments
2.2. Reagents, Plasmids and Media
2.3. Surface Marker Detection by FACS
2.4. Virus Production
2.5. Cell Lines
2.6. Differentiation of Immortalized HoxB8-FL Cells
2.7. DNA and RNA Isolation and Transcriptome Analysis
2.8. Short Tandem Repeat (STR)-Based Multiplex PCR
2.9. Fragment Length Analysis
3. Results
3.1. Differentiation and Characterization of Conditionally Immortalized HoxB8-FL Hematopoietic Progenitors
3.2. Authentication of HoxB8-FL Cell Lines by STR Profiling
3.3. Validation of Microfluidics-Based STR Profiling in Primary Murine Pancreatic Cancer Cell Lines
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Medium | Concentration | Media/Additive | Catalog No. | Storage |
---|---|---|---|---|
RP-10 Medium | RPMI1640 | Gibco: 21875-034 | 4 °C | |
10% | FBS (not heat inactivated) | Gibco: 10270-106 | −20 °C | |
0.10% | Mercapto Ethanol | Gibco: 31350-010 50 mM | 4 °C | |
1% | Antibiotic Antimycotic | Gibco: 15240-062 | −20 °C | |
Progenitor Outgrowth Medium | RPMI1640 | Gibco: 21875-034 | 4 °C | |
10% | FBS (not heat inactivated) | Gibco: 10270-106 | −20 °C | |
0.10% | Mercapto Ethanol | Gibco: 31350-010 50 mM | 4 °C | |
1% | Antibiotic Antimycotic | Gibco: 15240-062 | −20 °C | |
1 µM | β-estradiol | Sigma: E-2758 (stock 100 mM) | −20 °C | |
5% | FLT3L-SNT | from B16 melanoma cell line | −20 °C | |
B-Cell Differentiation Medium | RPMI1640 | Gibco: 21875-034 | 4 °C | |
10% | FBS (not heat inactivated) | Gibco: 10270-106 | −20 °C | |
0.10% | Mercapto Ethanol | Gibco: 31350-010 50 mM | 4 °C | |
1% | Antibiotic Antimycotic | Gibco: 15240-062 | −20 °C | |
1 µM | β-estradiol | Sigma: E-2758 (stock 100 mM) | −20 °C | |
5 ng/mL | FLT3L | R & D Systems: 427-FL | −20 °C | |
25 ng/mL | SCF | R & D Systems: 455-MC | −20 °C | |
7 ng/mL | IL-7 | R & D Systems: 407-ML | −20 °C | |
BBMM (Basal Bone Marrow Medium) | IMDM | Gibco: 12440-053 | 4 °C | |
30% | FBS (not heat inactivated) | Gibco: 10270-106 | −20 °C | |
0.20% | Mercapto Ethanol | Gibco: 31350-010 50 mM | 4 °C | |
0.50% | Antibiotic Antimycotic | Gibco: 15240-062 | −20 °C | |
1% | Glutamine | Corning: 25-005-CV | −20 °C | |
0.50% | BSA | Sigma-Aldrich: B6917 | 4 °C | |
HF2 Buffer | ddH2O | Ampuwa: 250166 | RT | |
30% | FBS (not heat inactivated) | Gibco: 10270-106 | −20 °C | |
0.50% | Antibiotic Antimycotic | Gibco: 15240-062 | −20 °C | |
1% | HEPES | Gibco: 15630-049 | 4 °C |
Cell Line | 18-3 | 4-2 | 6-7 | 9-2 | 15-3 | 6-4 | 12-1 | 5-5 | X-1 |
---|---|---|---|---|---|---|---|---|---|
#1_HoxB8FL (wild-type) | 16 | 19 | 19 | 17 | 22 | 19 | 15 | 16 | 23 |
#2_HoxB8FL (Iμ-HA-Bcl6) | 15 | 18 | 15, 18 | 15 | 20 | 18 | 16 | 17 | 24 |
#3_HoxB8FL (Cxcr4WHIM) | 16 | 19 | 19 | 15 | 22 | 19 | 16 | 17 | 24 |
Cell Line | 18-3 | 4-2 | 6-7 | 9-2 | 15-3 | 6-4 | 12-1 | 5-5 | X-1 |
---|---|---|---|---|---|---|---|---|---|
mPDAC06 | 17, 18 | 18 | 20 | 16 | 20, 22 | 17 | 16, 20 | 14, 16 | 20, 24 |
mPDAC09 | 17, 19 | 17, 19 | 20, 23 | 18, 22 | 20 | 18 | 16 | 16, 17 | 18, 25 |
mPDAC95 | 17 | 19, 19 | 17 | 18, 20 | 21 | 19 | 16 | 16, 17 | 26 |
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Hong, Y.; Singh, N.; Bamopoulos, S.; Gjerga, E.; Schmalbrock, L.K.; Balabanian, K.; Schick, M.; Keller, U.; Wirth, M. Authentication of Primary Murine Cell Lines by a Microfluidics-Based Lab-On-Chip System. Biomedicines 2020, 8, 590. https://doi.org/10.3390/biomedicines8120590
Hong Y, Singh N, Bamopoulos S, Gjerga E, Schmalbrock LK, Balabanian K, Schick M, Keller U, Wirth M. Authentication of Primary Murine Cell Lines by a Microfluidics-Based Lab-On-Chip System. Biomedicines. 2020; 8(12):590. https://doi.org/10.3390/biomedicines8120590
Chicago/Turabian StyleHong, Yingfen, Nikita Singh, Stefanos Bamopoulos, Enio Gjerga, Laura K. Schmalbrock, Karl Balabanian, Markus Schick, Ulrich Keller, and Matthias Wirth. 2020. "Authentication of Primary Murine Cell Lines by a Microfluidics-Based Lab-On-Chip System" Biomedicines 8, no. 12: 590. https://doi.org/10.3390/biomedicines8120590
APA StyleHong, Y., Singh, N., Bamopoulos, S., Gjerga, E., Schmalbrock, L. K., Balabanian, K., Schick, M., Keller, U., & Wirth, M. (2020). Authentication of Primary Murine Cell Lines by a Microfluidics-Based Lab-On-Chip System. Biomedicines, 8(12), 590. https://doi.org/10.3390/biomedicines8120590