Detection of in-situ hybridization to human metaphase chromosomes by atomic force microscopy

Naoaki Okamoto; Mitsuru Ishikawa

doi:10.1117/12.383354

21 April 2000 Detection of in-situ hybridization to human metaphase chromosomes by atomic force microscopy

Naoaki Okamoto, Mitsuru Ishikawa

Proceedings Volume 3922, Scanning and Force Microscopies for Biomedical Applications II; (2000) https://doi.org/10.1117/12.383354
Event: BiOS 2000 The International Symposium on Biomedical Optics, 2000, San Jose, CA, United States

Abstract

Detection of in situ hybridization to human metaphase chromosomes provides important information about gene mappings and about analysis of chromosomal disorders. We applied atomic force microscopy (AFM) to the detection of in situ hybridization to get better resolution as compared to light microscopy. Chromosomes were spread over a glass substrate and hybridized with DNA probes labeled with biotin or digoxigenin. The hybridized probes were reacted with streptavidin or anti-digoxigenin antibody, both of which were conjugated with 5-nm gold colloidal particles. We missed direct detection of the conjugated gold colloidal particles by micro-meter scale AFM scanning , but obtained clear topographic difference between the site of hybridization and the chromosome arm with the help of silver enhancement. We thus clearly detected the in situ hybridization using chromosome painting probes, alpha satellite probes, and locus specific gene probes by AFM. The in situ hybridization to DNA fiber was also detected by AFM. The detection of in situ hybridization by AFM has advantages over fluorescence in situ hybridization: no reduction of signal intensity under light irradiation. Application of AFM to the detection of in situ hybridization will be a useful method to analyze chromosomes.

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Naoaki Okamoto and Mitsuru Ishikawa "Detection of in-situ hybridization to human metaphase chromosomes by atomic force microscopy", Proc. SPIE 3922, Scanning and Force Microscopies for Biomedical Applications II, (21 April 2000); https://doi.org/10.1117/12.383354

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KEYWORDS

Particles

Gold

Atomic force microscopy

Latex

Silver

Microscopy

Luminescence

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