The Transient-State Polarization Fluorometer 1 (TSPF 1) is the first fluorometer which incorporates both patented Transient-State detection and Fluorescence Polarization measurement. The TSPF 1, exclusively from Diatron, offers the advantages of fluorescence polarization and "order of magnitude" increases in sensitivity from Transient-State detection in a single instrument.
Fluorescence Polarization (FP) used in a binding assay, allows the detection of the bound probe-target complex in a homogenous assay format. FP relies upon a measurable difference in polarized emission after excitation. The change in polarized emission is caused by a difference in speed of rotation which is a function of the size ratio of the bound to unbound probe.
The Fluorescence Polarization Binding Assay (FPBA) offers the advantage of performing the assay in a homogenous format, eliminating the need for separation of bound and unbound probe. This can eliminate the traditional assay washing steps, and may eliminate the need for isolation of the sample from a more complex fluid.
FP alone does not increase assay sensitivity (signal : noise) however, which is still affected by background noise from autofluorescence and scattered light (Raleigh-Raman Scatter).
The Transient-State detection measurement technique of our SIGNALL Technology does increase sensitivity by reducing the noise from both autofluorescence and Raleigh-Raman scatter.
Excitation is achieved by a series of short laser pulses. Photon counting is done in the intervals between pulses. The pulse from the diode decays very rapidly, unlike the decay from a lamp, and thus the scattered light is not registered.
The TSPF1 software is designed also to reduce the noise contribution from autofluorescence, further increasing sensitivity. As a result, 20 fold increases in sensitivity can be achieved compared to steady-state.
This increase in sensitivity expands the menu of tests which may be converted to fluorescence from the more cumbersome, expensive and hazardous formats, such as: radioactive (RIA), luminescent and colorimetric (ELISA), and greatly increases the opportunities for conversion to the homogenous FPBA format.
Transient-State should not be confused with the standard "time-resolved" methodology. Basic "time-resolved" detection normally cannot be used in fluorescence polarization assays and is limited to use with dyes of very long decay constants. Transient-State does not share these restrictions and limitations.