Single-cell detection of @entity470 gene mutation in circulating XXXX cells in @entity2 .

@entity1 @entity5 @entity470 @entity1149 @entity2 @entity3
UNASSIGNED: Circulating @entity5 cells (CTCs) are cells that shed from a primary @entity5 and enter the peripheral blood circulation. The CTCs are closely associated with @entity5 development and @entity3 because of its high heterogeneity. However, there are still no effective methods to detect single-cell heterogeneity of the CTCs. To this end, we developed a method to detect gene mutation in CTCs at the single-cell level and applied it to the detection of @entity470 gene mutation in @entity2 CTC. Specifically, the rare CTCs were captured from blood using an integrated microfluidic system, and then were released into a microchip with thousands of nanoliter wells to isolate single CTC. The single CTC was then transferred into a PCR tube under the microscope for single-cell genome amplification and detection of @entity470 gene mutation. We firstly modified chip and capillary and optimized PCR conditions (annealing temperature, number of cycles) using @entity1149 ( @entity1149 ) cell lines A549, NCI-H1650 and NCI-H1975 as samples, which showed maximal amplification after 30 cycles with an annealing temperature at 59   . We then successfully detected blood samples from @entity1149 @entity1 using this method. 5 CTCs were obtained from 2 mL @entity1 's blood and the sequencing of @entity470 exons 18, 19, 20 and 21 showed no mutations. Our results demonstrated that this method is sensitive enough to detect gene mutation in single CTC and has guiding significance in clinic research.
