Page 127 - The Indian Optician Digital Edition May-Junel 2022

MATERIAL & METHODS Table 2: Giemsa staining
Among the 50 cases included in our study, Fix smears in 15 minutes at least
30 were clinically diagnosed cases of trachoma, methanol
10 were of chronic conjunctivitis and 10 were Stain with diluted 20-30 minutes
normal controls. Conjunctival smears were giemsa
taken with a blunt spatula from each case and
spread on 2 slides for AO and Giemsa staining Wash with Sorenson’s buffer
respectively. Fix in Xylene-3 15 minutes each
changes
Dry and mount with DPX
Table 1: Acridine Orange (AO) Staining
Examine with light microscope
Fix smear in cold acid for at least 10 minutes
alcohol and Diluted Giemsa- Giemsa stain diluted with
Sorenson’s buffer in 1:2 dilution.
Wash with 3 changes 2 minutes each Sorenson’s buffer- Mixture of 50.80 ml. of 0.007
of Mc Buffer M Potassium dihydrogen phosphate KH2po4
Stain with AO 0.01% 6 minutes with 49.2 ml. of 0.07 M of Disodium hydrogen
(AO working solution) phosphate Na2 HPO4 at ph 6.8
Wash with 2 changes 3 minutes each
of Mc Ilvian Buffer TRACHOMA ??
Rinse with distilled water Blot and dry
Mount with buffer
Examine immediately with a fluorescent
microscope
Acid alcohol: absolute ethanol-2 parts glacial
acetic acid
Mc Ilvian buffer: mixture of 12.9 ml of 0.1 m citric
acid (21.015 gm of citric acid in 1 litre of distilled
water) and 7.1 ml of 0.2 M Disodium hydrogen
phosphate Na2HPo4 (28.393 gm of Na2HPo4 in
1 litre of distilled water at pH 3.8)
AO working solution: 1-10 dilution of 1% AO
stock solution with Mc Ilvian Buffer

RESULTS

The positive slides stained with AO were easily
diagnosed due to the presence of intracellular
fluorescent orange coloured elementary bodies
or green coloured initial bodies. The elementary
bodies are the infectious form and are
composed of mainly RNA. Initial bodies are the
non-infectious form and contain mainly DNA.
The positive slides stained with Giemsa showed
reddish blue elementary bodies or bluish purple STAGE 2
initial bodies in the slightly granular light blue TRACHOMA

CASE STUDY MAY-JUNE 2022 THE INDIAN OPTICIAN 123