Histological methods

Why this matters

You will send tissue to a laboratory from your first months in practice, and the pot you choose, the fluid you put in it and the mark you place on the specimen decide what a pathologist is able to report. Everything after that is done by people you will never meet, and every step of it changes the tissue: it shrinks, its lipid dissolves, its enamel disappears, and it is cut into slices thinner than the cells they contain. Reading a slide honestly means separating what belongs to the patient from what belongs to the process — and knowing that a mishandled specimen means your patient is re-biopsied.

The big picture

One biopsy becomes one slide through seven steps, and every step both reveals something and removes something. Fixation is diffusion, decalcification is a trade, sectioning is a slice thinner than the cell, staining is a chemical question put to the tissue, and the plane you look at was chosen before you ever saw the slide.

The 15 cards in this lesson

1. 01Four people make a slide, and exactly one step belongs to you
2. 02Seven steps, and the artefact each one leaves behind
3. 03Fixation has three clocks, and only the first one is diffusion
4. 04Clinical alert — in the lesson
5. 05Decalcification: the enamel is gone, and the agent decides what else you lose
6. 06The ground section: the preparation that keeps the enamel readable
7. 07Dehydration and clearing take the lipid with them
8. 08The specimen shrinks — mostly at the knife, and the numbers need checking
9. 09You never see a cell — you see a slice thinner than the cell
10. 10Haematoxylin is not a dye until you oxidise it and give it a metal
11. 11One stain answers one question — H&E is the default, not the answer
12. 12The light microscope stops at about 200 nanometres, and a high-power field is not an area
13. 13Electron microscopy: millimetre cubes, heavy metals, and no colour at all
14. 14The plane was chosen before you ever saw the slide
15. 15Artefact is the normal state of a slide — the skill is reading past it

You have got it when

You can (1) name the fixative for an oral biopsy, its volume and the liquids that do not fix tissue; (2) use d = 0.78 x root t to say when a block is fixed through; (3) say why enamel is absent from a decalcified section; (4) say what haematoxylin, eosin and PAS bind; (5) state the resolution limit of light microscopy; (6) tell an on-edge, en-face and tangential section apart; (7) state the TEM constraint and when…