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Q. What types of nucleic acid probes are frequently used?
A. Nucleic acid probes are complimentary sequences of nucleotide bases to the specific mRNA or DNA sequences of interest. They can be prepared in different ways. Currently, following types of probes are widely used:
- Oligo nucleotide probes - It is a single strand of a short DNA fragment generally ranging in size from 20-50 bases. The probes are chemically synthetic and generally end-labeled with hapten.
- Single stranded DNA probes - The probes are in the 200-500 base pairs in size range. They can be produced by reverse transcription of RNA or by amplified extension of a PCR-generated fragment in the presence of a single antisense primer.
- Double stranded DNA probes - These probes are generally produced by enzyme-based reaction using Nick translation reaction or PCR amplification
- RNA probes - The probes can be made by in vitro transcription. It is a single strand of RNA sequence.
Q. What are the nucleic acid probes labeled with?
A. Nucleic Acid Probes can be labeled with:
- Radioactive isotopes such as 35S, 32P and 3H
- Fluorochrome such as FITC, Texas Red, rhodamine, Cy5
- Hapten such as biotin, FITC, digoxigenin
- Enzymes such as an alkaline phosphatase
Different labeling needs different methods for downstream probe detection.
Q. What are the BioGenex probes?
A. The BioGenex probes are oligonucleotide (oligo) probes. They range in size from 20-40 bases. The probes are end-labeled with multiple fluoresceins. The fluorescein is a BioGenex proprietary product.
Q. How are the Biogenex oligo probes designed and validated?
A. Oligos are first selected from a given gene sequence by running a sophisticated computer software. Next, a blast search is conducted to check the homology of the selected oligos to human gene sequences in GenBank. Only target, gene-specific oligos are used. These oligos are synthesized and validated with multiple tissues, in different laboratories by two different technical experts at least.
Q. How to use BioGenex oligo probes?
A. BioGenex provides detection kits and a recommended protocol along with the oligo probes. Please see In Situ (ISH) Hybridization. If the BioGenex i6000 Automated Staining System and detection kits are used, the bar-coded system allows to conduct walk away automation, and generates publication-quality results within a day.
Q. What are the advantages of oligonucleotide probes over RNA or cDNA probes?
A. Advantage is the heritage of its smaller size compared to other probes. Following are main advantages for the oligo probes:
- More specific
- Very stable
- Easier to work with
- Better tissue penetration
- Faster to use
- High reproducibility
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