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Chapter 34 Calibration of Flow Cytometer Detector Systems

1994; Elsevier BV; Linguagem: Inglês

10.1016/s0091-679x(08)61097-3

0091-679X

Ralph E. Durand,

Single-cell and spatial transcriptomics

This chapter describes the calibration procedure of photomultiplier tube (PMT). The PMT calibration procedure requires ≤30 minutes for each filter-fluorochrome combination; the procedures are found to be sufficiently useful that express all quantitative measurements only in terms of calculated intensities. To facilitate these calculations further, it is useful to monitor PMT voltages and amplifier settings electronically in a manner that permits automated recording and use of these values by software packages. It should be obvious that the procedures allow standardization of fluorescence measurements, but do not substitute for the usual techniques of instrument alignment or for the necessity of internal controls in high-precision studies. These procedures do, however, permit quantization of fluorescence intensity over the entire usable range of the detectors, without analog-to-digital converter (ADC) or amplifier limitations. In view of the operational convenience of acquiring data with logarithmic amplification, the chapter also describes a method for conversion of logarithmic data to linear format.