PMT Gain Research Articles

Microarray scanner calibration curves: characteristics and implications

BackgroundMicroarray-based measurement of mRNA abundance assumes a linear relationship between the fluorescence intensity and the dye concentration. In reality, however, the calibration curve can be nonlinear.ResultsBy scanning a microarray scanner calibration slide containing known concentrations of fluorescent dyes under 18 PMT gains, we were able to evaluate the differences in calibration characteristics of Cy5 and Cy3. First, the calibration curve for the same dye under the same PMT gain is nonlinear at both the high and low intensity ends. Second, the degree of nonlinearity of the calibration curve depends on the PMT gain. Third, the two PMTs (for Cy5 and Cy3) behave differently even under the same gain. Fourth, the background intensity for the Cy3 channel is higher than that for the Cy5 channel. The impact of such characteristics on the accuracy and reproducibility of measured mRNA abundance and the calculated ratios was demonstrated. Combined with simulation results, we provided explanations to the existence of ratio underestimation, intensity-dependence of ratio bias, and anti-correlation of ratios in dye-swap replicates. We further demonstrated that although Lowess normalization effectively eliminates the intensity-dependence of ratio bias, the systematic deviation from true ratios largely remained. A method of calculating ratios based on concentrations estimated from the calibration curves was proposed for correcting ratio bias.ConclusionIt is preferable to scan microarray slides at fixed, optimal gain settings under which the linearity between concentration and intensity is maximized. Although normalization methods improve reproducibility of microarray measurements, they appear less effective in improving accuracy.

TU-A-I-617-02: PET Scanner Performance: Quality Assurance and Acceptance Testing

Quality assurance of PETscanners must be performed on a regular basis to maintain and confirm proper scanner performance. These procedures should track system stability and be sensitive to changes in scanner operation. The quality control and calibration of a PETscanner includes detector and electronic characterizations such as adjustment of PMT gain, definition of crystal and energy maps and coincidence timing calibration. Briefly, the crystal map converts the analog position of the detected event to a specific crystal within the block detector, while the coincidence timing calibration adjusts for timing delays so events from each block are time stamped equivalently. These characterizations are applied to the PET data during acquisition. Daily quality assurance should determine whether the scanner is suitable for clinical use. A uniform cylinder of activity or rod sources are a good source of quality control data. These scans can be used to monitor system stability and to determine which crystals, blocks and modules (buckets) are more (or less) sensitive than the respective system average. The NEMA PET NU2‐2001 standard should be followed for acceptance testing. This standard uses a polyethylene phantom of 700mm axial length with a 3.2 mm line source to measure scatter fraction, count losses and randoms. This phantom approximates the out‐of‐field activity distributions of whole body scans. The measurement of sensitivity is conducted with a line source surrounded by known absorbers, and the sensitivity with no absorbers can be found by extrapolation. The intent of the image quality measurement is mimic a whole body scan using a torso phantom containing hot and cold spheres of various diameters (representing lesions) in a warm background. This presentation will focus on the calibrations and corrections required to maintain proper system performance. The presentation will also describe the rationale and methodology of the NEMA NU2‐2001 performance standards.

The MINOS light-injection calibration system

A description is given of the light-injection calibration system that has been developed for the MINOS long-baseline neutrino oscillation experiment. The system is based upon pulsed blue LEDs monitored by PIN photodiodes. It is designed to measure non-linearities in the PMT gain curves, as well as monitoring any drifts in PMT gain, at the 1% level.

Calibrations and corrections of a modular cylindrical SPECT system (McSPECT-II)

The authors have developed a modular cylindrical SPECT system (McSPECT-II) for clinical brain imaging. This system features a stationary cylindrical detector and a rotating collimator. The cylindrical detector is comprised of 120 NaI(Tl) crystal bars coupled to a circular array of PMTs. The output signal from each PMT is digitized by an ADC. The position calculation and corrections are performed using software and precalculated lookup tables on a Pentium-based PC. The collimator has four quadrants, and each quadrant consists of a parallel array of 25 collimator units. The four corner wedges between adjacent quadrants are utilized to accommodate line sources for system calibrations. The authors report the calibration and correction procedures developed for the McSPECT-II system. These procedures include: (1) PMT gain normalization, (2) azimuth bar identification, (3) energy correction, (4) axial linearity correction, and (5) uniformity correction. Results of these calibrations and corrections on data acquired by imaging various phantoms are presented.

YAlO 3: Ce-Am light pulsers as a gain monitor for undoped CsI detectors in a magnetic field

YAlO 3-Am pulsers show no degradation in their light output or resolution upon the application of a magnetic field, at least up to 10 kG. The decrease in the pulse height agrees with the change in the PMT gain calibrated by an LED. When the pulser is mounted on the top face of a 30 cm long CsI far from a PMT with a UV filter inserted between the PMT and the CsI, the light output decreases by a factor of 1 4 , compared with the case of being directly mounted on the PMT window. In the above configuration, the 5.49 MeV α of 241Am gives about 550 photoelectrons, which corresponds to a γ-equivalent energy of 6.6 MeV. Finally, the pulsers were mass-produced with a FWHM resolution of 5.4% (average) for the pulse height.

Novel method of measuring optical-intensity distributions of scintillations

A new method of measuring optical-intensity distributions of weak scintillations is described that uses the photon counting technique in combination with a variable optical attenuator. It is experimentally demonstrated that the proposed method permits the distributions to be determined from a set of integral data of count rates measured at different optical-attenuation factors. In addition to being almost insensitive to possible changes in PMT gain, the present method offers the advantage over the conventional one relying on pulse height analysis that it can provide virtually improved energy resolutions in measuring low-intensity scintillation energy spectra.

Gated photomultiplier response characterization for DIAL measurements

The characteristics of various detector responses are studied to understand the cause of various systematic biases and to minimize these undesirable effects in measurements of transient signals with large dynamic range. We quantitatively evaluated signal induced bias, gain variation, and the linearity of commonly used gated photomultipliers in the current integrating mode. Analysis of the results indicates that impurity ions inside the photomultiplier tube are the source of the signal induced bias and gain variation. Two different photomultiplier tubes used in this study show significant differences in the magnitude and decay behavior of signal induced bias. We found it can be minimized by using an external amplifier to reduce PMT gain, and by applying a low potential between the cathode and first dynode. The linearity of a photomultiplier tube is also studied over a large dynamic range of input intensities employing a new technique which does not require an absolute calibration. The result of this study shows that the photomultiplier response is linear only for a limited input intensity range below a certain anode current.

An Automated System for Energy-Loss Microanalysis

Figure 1 shows a system which we are using for storage and display of electron energy-loss spectra, based on a Tracor Northern TN-1710 multichannel analyser. For serial acquisition, scan coils of the magnetic spectrometer1 are driven directly from the ramp output of the multichannel analyser. The electron detector consists of a transmission phosphor followed by a photo-multiplier tube, whose analogue output is fed via a 5 MHz voltage-to-frequency converter (Teledyne model 4707) into a TN-1710-26 plug-in scaling module. To allow adequate dynamic range (>108) and avoid overloading the photomultiplier tube, PMT gain is changed by a factor G ˜ 500 during acquisition. This is done automatically by incorporating into the line from the power supply (Venus model LR-2N) a series of Zener diodes which are short-circuited by a reed relay (driven from a high-gain amplifier) when the scan voltage exceeds a preset threshold (fig. 2). The gain change is accomplished in a shorter time (a few ms) than was possible by voltage-programming the power supply, which makes the factor G easier to measure accurately.