Study Of The System Design And Methods For Measuring Performance Of The Single-laser Four-channel Flow Cytometer

So far, flow cytometry assay is widely used in scientific research and moderntherapy. This advanced technique is consisted of physical electronics, laser andcomputer technology, and can be used for the multi-parameter detection of physical,chemical and biological characteristics for tens of thousands of cells or biologicalparticles within seconds. Meanwhile, it is of a high detection rate, accuracy andsensitivity. Flow cytometer is an instrument with flow cytometry assay as the coretechnology, and is called as "CT" for its important applications and far-reachingeffect on our scientific research and ordinary lives. Unfortunately, no flowcytometers have been produced in China, indicating a desire for the furtherdevelopment of flow cytometry assay. In our work, flow cytometry is studiedintensively from the designning, debugging and quality evaluation of the verificationsystem; chicken red blood cell preparation process, instrument quality control andsteatosis; size differentiation and absolute quantification of micron particle; thestructure simulation design of the sorting flow chamber.1. In accordance with the characteristics of flow analysis, the choice ofexcitation source and detection region, the design of the excitation optical path,spectral separating systems, flow-driven system and fluorescence detector modulesare carried out. A verification system of the flow cytometer is built with a532nm laser as the excitation source, the flow chamber of the integrated optical system andavalanche diode detector modules. The entire system achieves the absolute count ofcells; simple debugging of the optical path, the detection of the weak fluorescence.Then, spherotech rainbow calibration particle RCP-30-5A is used to evaluate theperformance of the system, and the fluorescence signal histogram of RCP-30-5Amicrosphere shows that the number of individual peaks is eight, dynamic range is105, system sensitivity is76MEPE, Q=0.087, B=374MEPE. The analysis of thequality curve and the scientific, reasonable quality assessment system are built.2. Calibration particles for routine calibration of flow cytometers are importedand expensive, and the cost of maintenance and debugging increases for thesereasons. The trial chicken erythrocytes for flow cytometry cost low and meet thequality control requirements of the debugging, and the method of the quality controlis set up on them. Combined with the flexibility of the build system, it is possible toexpand the application of the system. At last, the steatosis is investigated onverification system.3. Rapid, quantitative count and size differentiation analysis of micro-sizedpolystyrene microspheres are done via single particle detection, which completes thestatistical analysis, achieves micro-sized particles count and paves the way for theapplication of biomedical and environmental monitoring at the same time.4. The flow chamber as a significant role in flow cytometry is the corecomponent of flow cytometer. Simulation of the flow chamber gives the focus of thesheath flow and sample flow. Using particle tracking analysis, simulation of the jetperturbation of the sample flow for sorting the cell provides a reference to the sortingflow chamber design.