1.0 Purpose
This SOP establishes a standardized protocol for Giemsa staining of cytology smears and tissue sections.
2.0 Scope
This procedure is applicable to all cytology smears and tissue sections intended for Giemsa staining within the Surgical Pathology section of the laboratory.
3.0
Introduction
Giemsa staining is utilized for evaluating non-gynecological smears from Fine Needle Aspiration (FNA), body fluids, and tissue sections, particularly for detecting parasites.
4.0
Principle
Giemsa staining involves a differential process utilizing Azure II/Eosin. This stain's action relies on the interaction of these compounds. Eosin stains structures with a more basic nature in red, providing clear cytoplasmic details compared to Papanicolaou (PAP) stain.
5.0
Analytical Measuring Range and Limit of Detection
Not applicable.
6.0
Responsibility
This SOP is the responsibility of both technical staff and pathologists.
7.0
Abbreviations
NIL (No abbreviations used).
8.0
Requirements
Reagents:
(a)
Distilled water
(b)
Absolute methanol
(c) Giemsa
stain solution
(d)
Immersion oil
(e) DPX
Instrumentation
and Software:
(a) Slides
(b) Slide
rack
(c)
Pasteur pipettes
(d) Light
microscope
(e) Cover slips
Disposables:
NIL
Other
Materials required but not provided with the kit (Reagents and Consumables):
NIL
9.0
Precautions
All specimens and instruments must be handled with gloves, masks, and goggles to ensure safety and prevent contamination or injury.
10.0
Limitations & Interferences
Not applicable.
11.0
Documentation
All relevant documents are maintained in the Daily Routine Register and Special Stains Log/Register.
12.0
Instructions
12.1
Pre-Analytical Steps:
12.1.1
Giemsa Stain Preparation:
12.1.1.1 Giemsa Stock Solution:
Rinse a
clean bottle with methanol.
Place
clean and dry glass beads in the bottle.
Add 62.5
ml of glycerol and 1 gm of Giemsa Stain Powder.
Mix well
and incubate at 60°C with intermittent shaking for 1 hour.
Add 62.5
ml of methanol and allow to stand for 2 to 7 days before filtering before use.
12.1.2
Giemsa Working Solution:
Mix 1 ml
of Giemsa Stain stock Solution with 9 ml of Buffered Water (pH 6.8) just before
use.
12.2 Analytical Steps
Mark the
cytology slide with a diamond pencil.
Fix smear
for 15-20 minutes in absolute methanol.
For tissue
sections, remove wax with xylene and alcohol, then pour diluted Giemsa working
solution over the sample.
Allow
stain to act for 20 to 25 minutes.
Wash with
distilled water and dry slide upright.
Mount
smears directly; dehydrate tissue, clear, mount, and label.
Present
slides to pathologist for reporting.
12.3
Quality Control and Assurance
Run a known positive control slide with each batch of samples.
12.4
Calibration and Calibration Verification
Not applicable.
12.5
Instructions
Storage of
Samples:
Store slides for 10 years.
13.0 Reference
Range
Not applicable.
14.0
Critical Value
Not applicable.
15.0
Reporting of Results and Interpretation
Results are reported based on the morphology of organisms/cells being evaluated.
16.0
Contingency Plan
Multiple trained staff and backups are available in case of reagent shortages, manpower issues, or instrument breakdowns.
17.0
References
Bancroft:
"Theory and Practice of Histological Techniques" by John D Bancroft
and Marilyn Gamble, 2007 (6th ed).
END OF THE DOCUMENT
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