Histological Technique Overview

Histopathology Overview

• Histopathological Technique studies tissue structures in diseases.
• Specimens originate from surgeries, biopsies, and autopsies in various forms.

Tissue Preparation Steps

• Receiving, grossing, decalcification (if necessary), fixation, dehydration, clearing, impregnation, blocking, section cutting, staining, and mounting are critical processes for microscopy.

Specimen Receiving Room

• Ensures safe and secure intake of samples.
• Biopsy specimens must be checked for fixatives, tissue presence, patient info, and physical attributes.

Steps for Receiving Biopsy Specimens

• Verify presence of fixative and tissue.
• Document patient details (name, age, sex, diagnosis) in a clinical report book.
• Proper labeling of specimens is essential to avoid diagnosis errors.

Criteria for Rejecting Specimens

• Inconsistent patient details between requisition and specimen labels.
• Unspecified site/nature of specimen.
• Incorrect containers or lack of appropriate fixatives.
• Missing patient history or essential content in blocks/slides.

Types of Specimens

• Surgical Biopsy, Needle Biopsy, Endoscopy Biopsy, Incision Biopsy, Excision Biopsy, Punch Biopsy, Cone Biopsy.

Grossing Techniques

• Grossing involves inspecting, measuring, and sectioning tissue samples.
• Ideal size for processing is approximately 20mm x 15mm x 5mm.

Requirements for Grossing

• Necessary tools include various scissors, forceps, scalpels, weighing scales, cassettes, and disinfectants.

Decalcification Process

• Essential for removing calcium ions from calcified tissues to allow proper sectioning.
• Must avoid damage to tissue cells and fibers.

Good Decalcifying Agents Characteristics

• Must ensure complete calcium removal without damaging tissue.
• Should not impair staining and offer reasonable decalcification speed.

Decalcification Procedure

• Involves selection, fixation, decalcification, neutralization of acids, and thorough washing.

Fixation Overview

• Fixation preserves tissue morphology; critical for accurate histopathological preparation.
• Tissues must be fixed immediately post-surgery to prevent autolysis or putrefaction.

Fixative Quantity and Process

• Fixative volume must be 15-20 times that of the tissue; fixation takes 12-24 hours at room temperature.

Purposes of Fixation

• Prevents autolysis and putrefaction.
• Preserves tissue for future studies and enhances staining.

Common Fixatives

• Aldehydes (e.g., formaldehyde), protein denaturants (e.g., alcohol), oxidizing agents (e.g., osmium tetroxide).
• Various fixation techniques include immersion, vapor, microwave, and perfusion.

Specific Fixatives and Their Characteristics

• Formalin (10%): Cheap, preserves most tissues well, but can be allergenic.
• Zenker’s Fluid: Excellent nuclear preservation, but penetrates minimally and is expensive.
• Bouin’s Fluid: Rapid penetration; preserves glycogen but risks red cell lysis if overexposed.
• Helly’s Fluid: Preserves nuclear structures, facilitating staining but has limited penetration.

Overview of Additional Fixatives

• Baker’s Solution, Orth's fluid, Carnoy’s fluid also used in various scenarios for tissue fixation.