Laboratory Equipment Blog
The use of fluorescence for sensing and imaging of the cellular signaling pathways has emerged as an indispensable tool in modern physiology, providing dynamic information of quantity and localization of the molecules of interest. Using appropriate indicator dyes, molecules alter their fluorescent characteristics in response to ion binding or membrane integration, so that the optical signal from the indicator can be measured to monitor the amplitude and the time course of various metal ions like Na+, K+, Mg2+ and Ca2+, as well as pH and membrane potential, in cellular compartments.
Absorption of light correlates to the energy of a photon that is taken-up by electrons of the substance atom. The electromagnetic energy is transformed into internal energy of the absorbent substance. The absorbance of a substance quantifies how much of the incident light is absorbed by it (instead of being reflected or refracted). Precise measurements of the absorbance at many wavelengths allow the identification of a substance via absorption spectroscopy, where a sample is illuminated from one side, and the intensity of the light that exits from the sample in every direction is measured (see Fig. 1). A few examples of absorption are ultraviolet–visible (UV-Vis) spectroscopy or infrared (IR) spectroscopy.
The use of fluorescent probes in cell physiology has emerged as indispensable tool in the analysis of cell functioning over recent years. The physics underlying fluorescence is illustrated by the electronic-state diagram (so-called Jablonski diagram, see Fig. 1), showing the three-stage process to create the fluorescent signal (Excitation - Excited/State Lifetime - Fluorescence Emission) in a fluorophore/indicator and simplified described below.
Concentrations of DNA in solution (31µg/mL and 144µg/mL) were measured with a spectrometer and UV/VIS light source in a SPT-2. Due to the 1cm pathlength, use of a SPT-2 does not require a pre-measurement dilution within this concentration range, thus a potential source of error was eliminated. Due to the design of the SPT-2, routine measurements can be taken with samples of 10µL in standard 200µL PCR tubes. If caution is taken with regard to the location of the probe tip within a PCR via, measurements remain repeatable in sample sizes as small as 5µL.
Concentrations of DNA in solution (31µg/mL and 688µg/mL) were measured with a spectrometer and UV/VIS light source in a DIPUV-Mini. Due to the 2mm pathlength, use of a DIPUV-Mini does not require a pre-measurement dilution within this concentration range, thus a potential source of error was eliminated.
Concentrations of DNA in solution (31µg/mL and 561µg/mL) were measured with a spectrometer and UV/Vis light source in a cuvette. A 2mm pathlength cuvette does not require a pre-measurement dilution within this concentration range, thus a potential source of error was eliminated. Although a 2mm cuvette has a total internal volume of 0.7mL, only 350µL is required to obtain an accurate measurement.
WPI's LEDspec is an LED-based, multi-wavelength detector with integrated reference channel. LEDspec is suitable for a variety of applications from nutrient analysis to water purity and trace metal detection.
Cuvettes come in a variety of shapes and sizes, but one of the most important specifications of a cuvette is its Z-dimension. The Z-dimension of an instrument (cuvette holder or spectrometer) is the distance from the bottom of the cuvette chamber floor to the center of its light beam (see image). A cuvette’s Z-dimension must match the Z-dimension of the instrument with which it will be used.