How To Examine Cells And Tissues Flashcards
Epithelial tissue
Often on the edge of other tissues and surround other tissues
Sometimes in clusters within other tissues (glands)
Polarised when at surfaces
Always have a basement membrane on the basal (lower surface)
Often secrete something from apical surfaces
Held together by strong anchoring proteins
Communicate through ‘junctions’ at their lateral and basal surfaces
Sugar on surface of cells - glycocalyx - attracts water - therefore keeps us hydrated
Connective tissue
Consist of cells and extracellular proteins/ glycoproteins and ‘gels’
Main cells are:
Fibroblasts - ligaments
Chondrocytes - cartilage
Osteocytes/osteoblasts/osteoclasts - bone
Stem cells/progenitor cells/bone marrow/blood/adipocytes
Main products are:
Fibres (many different types)
‘Ground substance’
Wax and gel-like materials
Nerve tissue
Nerve cells (neurons) and several support cells
Nerves can be relatively short (μm)
Can be very long (cm)
Main fast communication system in the body
Cells congregate into nerve fibres
Fibres congregate into ‘nerves’ that can be dissected and visualised ‘by eye’
Sciatic nerve is longest nerve in the body running down back of leg
Muscle tissue
Consists of muscle cells
3 types
Skeletal Cardiac Smooth
Major Function: to contract!
Enables Movement (the organism)
Provides Stability (the organism, organs and tissues)
Allows for Movement of tissue contents - via peristalsis
Minor Function: to secrete hormones
e.g. Natriuretic factor(s)
Myostatin(s) - stops heart from growing too big
Comparison of light microscopy vs electron microscopy
Light microscope: Can view images in natural colours Large field of view Cheap and easy preparation Can view living and moving objects However - low magnification and low resolution
Electron microscope High magnification High resolution However - only monochrome images can be see Have a limited field of view Difficult and expensive preparation Can only view dead and inert objects
Freeze fracture Electron microscopy
The tissue is frozen to -160oC and fractured by hitting with a knife edge.
The fracture line passes through the plasma membrane exposing its interior which can then be imaged.
Gives you an indication on how many transmembrane proteins there are in a proximal tubule
Paraffin wax formalin fixed vs frozen section
Paraffin embedded tissue selection - these come from preserved fixed tissues that take around 24-48 hrs to make - they are permanent and very useful for pathological diagnosis
Frozen section - taken from fresh tissues, takes around 10-20 minutes to make, only last for a couple of months and are usually used for emergency diagnosis in an operation to check to see if something is cancerous
Immunochemistry
1o antibody binds to the foreign protein, the 2o antibody has a fluorescent tag/enzyme attached, therefore can be detected with light/staining, indicating that the cell is infected (immunofluorescence)
Confocal microscopy
Laser excites a fluorescent molecule and electrons in the dye are raised to higher energy level
As the electron relaxes back to its ground state (lower energy level) a light with higher wavelength emitted
Emitted light is sent through mirrors and a pinhole screen to a CMOS detector (like the one in your mobile phone’s camera)
Because only one part of light (that bit that’s in focus) reaches the
detector, the images are very sharp
Means the entire depth of the cell/tissue can be examined
Motorisation allows full section scanning
Preparation of live cells
Process to culture cells -
Harvest cells
Isolate cells with the use of collagenase and DNAse
Apply the isolated cells on to an appropriate growth media (culture dish)
Subculture / centrifuge cells to obtain a pure sample for the culture
Verify the cultured cells are the cell type of interest
Cells then can be manipulated or modified for experimental procedures
What needs to be maintained constant in the internal environment - Concentration of oxygen, carbon dioxide, salt and other electrolytes
- Concentration of nutrients, waste products
- pH of internal environment
- Temperature of internal environment
- Volume and pressure of fluid and cell compartments equalised
Cells in culture
Cells in culture • Allows manipulation of the cells and experiments to determine cells and thus tissue function
Advantages/disadvantages of Cell culture
Advantages:
Absolute control over the physical environment
Homogeneity of sample
Less need for animal models
Disadvantages:
Hard to maintain
Only grow small amount of tissue at high cost
Dedifferentiation can occur
Instability, aneuploidy
3 dimensional architecture is lost
Influence of other cells/tissues not maintained
