title: Microdensitometric scanning procedure for quantitative assessment of hybridization of rRNA targeting probes in leprosy.
abstract: In order to develop an objective criteria of grading of positivity of hybridization signals of gene probes targeting rRNA, a microdensitometric scanning procedure was standardised. Ribosomal RNA was extracted from the bacilli harvested from biopsies of leprosy cases across the spectrum and blotted on nitro-cellulose membranes. M. leprae specific rRNA targeting oligonucleotide probes were end-labelled and hybridization was done by the technique standardised and published earlier. The autoradiographs were developed and microdensitometric scanning was done by altering different parameters. Positivity was graded in 5 grades and compared with visual positivity. Microdensitometric scanning procedure and 5 grade system appear to be useful and reproducible. Signals in paucibacillary specimens were in 2+ to 3+ grading range whereas those in multibacillary specimens varied in grades from 2+ to 5+. This approach appears to have potential usefulness for assessing the bacillary load (possibly viable) in the clinical specimens from leprosy cases.
authors.affiliation: Central JALMA Institute for Leprosy (ICRM), Tajganj, Agra, India.
authors.forename: R K
authors.initials: RK
authors.lastname: Sharma
authors.forename: C T
authors.initials: CT
authors.lastname: Shirannavar
authors.forename: K
authors.initials: K
authors.lastname: Katoch
authors.forename: V D
authors.initials: VD
authors.lastname: Sharma
authors.forename: M
authors.initials: M
authors.lastname: Natrajan
authors.forename: N
authors.initials: N
authors.lastname: Saxena
authors.forename: V M
authors.initials: VM
authors.lastname: Katoch
mesh_terms: D001345:Autoradiography; D001706:Biopsy; D015152:Blotting, Northern / Q000379:methods*; D003720:Densitometry / Q000379:methods*; D006801:Humans; D007918:Leprosy / Q000382:microbiology* / Q000473:pathology; D009166:Mycobacterium leprae / Q000235:genetics / Q000302:isolation & purification*; D015345:Oligonucleotide Probes; D012329:RNA, Bacterial / Q000032:analysis*; D012335:RNA, Ribosomal / Q000032:analysis*; D015203:Reproducibility of Results
chemical_list: D015345:Oligonucleotide Probes; D012329:RNA, Bacterial; D012335:RNA, Ribosomal