A microarray is an instrument that helps to detect the expression of thousands of genes at the same time. The microarray is called gene chips or DNA (deoxyribonucleic acid) chips because the slide consists of tiny spots for embedding DNA like the microchip. The DNA molecules in each slide act as a probe for detecting gene expression. The probe DNA molecule is also called the transcriptome.
Probe DNA are single stranded DNA that helps to detect target complementary DNA by hybridization.
Microarray scanners are laboratory equipment that helps to measure the fluorescent areas of DNA microarray, revealing information about the activity of thousands of genes. It helps to determine the quality of gene expression or overexpression of genes during particular disease.
Procedure of Microarray
The basic principle of DNA microarray is “nucleic acid hybridization.” Nucleic acid hybridization means joining two complementary DNA by hydrogen bonds to form a double-stranded molecule. The procedure of microarray has the following steps:
Preparation of DNA chip
There are many ways to prepare a microarray chip. The DNA chip was designed using photolithographic methods in the early days. Nowadays, photoactivated chemistry and masking help obtain DNA probes which are also available for purchase commercially. Also another commercially available way is an already designed probe attached to a fine needle printed on a chemical matrix surface by a robot.
Nucleic acid hybridization
For hybridizing the nucleic acid, the following steps are performed:
- Sample collection: Two samples of healthy as well as infected tissue for comparison are preferred.
- Isolating RNA (ribonucleic acid): Isolate mRNA from the collected sample using the column or solvent method.
- Labeled cDNA preparation: Prepare labeled cDNA (complementary DNA) from the isolated RNAs. Labeling is done by using fluorescent dyes like Cy3 and Cy5.
- Hybridization: Now, place the prepared, labeled cDNAs into the DNA chips/microarray trays with the required probe for hybridization.
Analysis of DNA chip
A microarray scanner helps collect the data of thus hybridized chip/microarray tray after the tray is rinsed thoroughly to remove the unbound/unhybridized DNAs. The intensity of the color is measured, and the difference helps analyze the genes.
Principle of Microarray Scanner
The lasers of the microarray scanner excite the fluorescent-labeled genes in the microarray tray. Likewise, the camera and microscope help form the image of excited genes. Then a program helps calculate the red to green ratio or subtract background data for each spot in the microarray using the digitalized image of the hybridized gene.
Parts of Microarray Scanner
The parts of a microarray scanner are broadly classified into two parts:
- Scanner: It consists of the laser for exciting the labeled hybridized DNA and a photomultiplier tube for reading each chip spot one by one.
- Imager: It consists of a laser for exciting the DNA, a camera and a microscope for capturing the image of the gene.
- Analyzer: It is a software program that helps analyze the data generated after scanning by comparing the color contrast.
Benefits of Microarray
Analysis of multiple samples
Since a single chip consists of thousands of spots for DNA hybridization, a single chip can analyze multiple samples simultaneously. It is also beneficial in separating the non-cancerous cells from the cancerous cells.
The microarray technique helps produce consistent data, which increases the speed of analysis in comparison to other nucleic acid hybridization techniques.
Compared to conventional assays, the area or scale used in the microarray is minimal, reducing reagents, minimizing research volumes, and increasing the speed of reaction and sample concentrations.
Limitation of Microarray
The result generated by microarray is usually qualitative; that is, quantifying the data obtained is impossible.
Even though the used probe is the same, sometimes the result can differ from the original one. Similarly, posttranscriptional monitoring can affect the translation.
Not applicable for proteins and lipids
Most gene expression techniques have been modified to accommodate the detection of end products like proteins and lipids. Still, microarray only deals with the detection of targeted genes/DNA.
How to Choose a Microarray Scanner
The answer to choosing a microarray scanner fit for your laboratory is checking all the following requirements:
- Sensitivity: The microarray scanner’s sensitivity must be high to avoid any DNA analysis errors.
- Resolution: The microarray scanner must have a camera with good resolution to magnify the microscopic image.
- Scan area: The scan area of microarray depends on the number of spots usually analyzed in your laboratory. Possibly a slightly larger area than the spots generally analyzed because the number of samples to analyze might increase in the future.
- Scanning speed: The scanning speed highly determines the correct microarray scanner for your laboratory. You should prefer a microarray scanner with the highest scanning speed because it determines the speed to completion of the microarray process.
- Other considerations: Other considerations like how easy is it to use, do you require multiple laser and filter options, and whether it is upgradable according to future needs are also necessary.
Surescan Dx by Agilent Technologies and GeneChip 3000 7G by ThermoFisher Scientific fulfill the basic requirements of a microarray scanner.
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