Microscopical analysis BIOL30002 Experimental Reproductive Physiology BIOL30002 Experimental Reproductive Physiology Techniques: Microscopy Microscopical analysis Structure   Function Gross anatomy (macroscopic) tells us what is happening on a gross scale Microscopical anatomy tells us what is happening on a cellular scale

Microscopy Scale limited by detection method visible light: wavelength ~ 0.38 – 0.74 μm  cannot reveal structures less than about 1 μm (organelles) Electron beam: at 10kV  0.00 001 μm (large molecules) Synchrotron beam:  0.000 001 μm (small molecules) Atomic force microscopy  atomic scale

Types of microscopy Light microscopy thin sections – paraffin; frozen – 4-10 μm good for tissue/cellular level relatively low cost many ways to stain to highlight things of interest Confocal microscopy light microscopy with special optics can get higher resolution can build up 3-D structure Transmission Electron microscopy extremely thin sections (0.05 μm) good for cellular and subcellular level expensive machinery more limited staining options (heavy metals) Scanning EM shows surface structures

Interpreting Microscope Slides 2-D vs

histology light microscopy vs TEM vs SEM vs confocal stains Resources: haematoxylin & eosin - good general purpose trichrome - good connective tissue discrimination PAS - sugar containing components lipophilic dyes - fats and oils nucleic acid stains enzyme histochemistry immunohistochemistry Resources: Kerr (2000) Atlas of Functional Histology Wheater et al, (1987) Functional Histology

Basic tissue types blood connective tissue, bone fat (adipose tissue) muscle nerves epithelium (epithelia)

epithelia columnar cuboidal squamous … lumen Junctions between cells  barrier columnar cuboidal squamous … lumen uterine luminal epithelium

pituitary stalk (with blood vessels) posterior (nerve ends) anterior

testis epididymis testis fat epididymis

BIOL30002 Experimental Reproductive Physiology Techniques: Microscopy seminiferous tubules lumen blood germ cells

BIOL30002 Experimental Reproductive Physiology ovary Techniques: Microscopy antral follicle oocyte

Drawings Use a SHARP pencil Tissue outlines – don’t attempt elaborate shading unless you are very skilled Use appropriate size scales Don’t spend forever replicating fine detail – just do a bit to show Label and annotate with observations and info from reference material