What Is a Microspectrophotometer?
A microspectrophotometer is a spectrophotometer specialized for the measurement of very small amounts of samples.
Like the spectrophotometer, it is intended for quantitative analysis to determine the concentration of specific substances in a sample. It can analyze the amount of DNA, protein, and other substances contained in a sample, and is widely used in the life science field.
The instruments themselves are compact and easy to handle, and models with touch panels and models that can easily output data externally are also available.
Applications of Microspectrophotometers
The main application of microspectrophotometers is the determination of DNA, RNA, oligonucleic acids, and proteins from extremely small amounts of samples. They are mainly used in research and development in the life science field.
The quantification and purity evaluation of nucleic acids and proteins is a necessary process in genomic DNA preparation, RT-qPCR, and sample quality control. Kinetic evaluation is also performed by monitoring bacterial culture growth and measuring over time.
Principles of Microspectrophotometers
The basic principle of the microspectrophotometer is the same as that of the Spectrophotometer. The spectrophotometer quantifies the concentration of a particular substance in a sample based on how much light shined on the sample reaches through it. The higher the concentration, the lower the transmission because light is blocked by substances dissolved in the sample.
Structure of Microspectrophotometers
A spectrophotometer is an instrument consisting of a light source, a sample cell (such as a sample port in the case of a microspectrophotometer), and a detector. The light emitted from the light source is divided into monochromatic light using a monochromator, irradiated onto the sample, and the light that passes through the sample (transmitted light) is measured using a detector.
A calibration curve of the amount of transmitted light (absorbance) and concentration is prepared using a solution of known concentration to determine the concentration of the unknown sample.
Types of Microspectrophotometers
Microspectrophotometers are generally capable of measuring very small sample volumes, but the minimum volume varies from instrument to instrument. For example, there are 0.3 µL, 0.5 µL, 1 µL, etc. It is important to select the appropriate one for the sample to be used.
On the other hand, some instruments can be used in a cell or on a cuvette board to measure samples of a certain volume. Some microspectrophotometers are also equipped with algorithms and programs to help perform advanced analyses.
For example, those equipped with programs that allow measurements over time at specific wavelengths are useful for performing kinetic analysis. Others are equipped with a reference library of spectra and impurity prediction algorithms that can predict impurities in a sample and calculate more accurate sample concentrations.
Other Information on Microspectrophotometers
1. Sample Handling for Microspectrophotometer
In an ordinary spectrophotometer, the sample is placed in a cell for measurement. In a microspectrophotometer, however, the sample is so small that it is generally dropped into the sample port with a pipette for measurement. Some instruments can be used for measurements other than micro amounts, and it is possible to analyze samples by placing them in a cell as in a regular microspectrophotometer, or by using a cuvette board.
In microspectrophotometers, it is often important that droplets are formed by surface tension, so each product has its mechanism to ensure proper measurement. Some instruments are also designed to confine the sample in a sealed space to reduce the risk of evaporation or contamination.
2. Light Source
Xenon flash lamps are often used as the light source. Xenon flash lamps are characterized by high brightness and low heat generation from the lamps. This reduces damage to samples that are sensitive to temperature changes.