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OSSF > OSSF-Histology techniques > Flashcards

OSSF-Histology techniques Flashcards

1
Q

What are the different techniques used to study tissue microscopic anatomy?

A

Light microscopy, cytology, scanning electron microspy, transmission electron microscopy, histochemical stains, frozen section, immunohistochemisrtry techniques.

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2
Q

what are the steps used to process tissue histology/ histopathologic examinations (tissue biopsy)?

A

-FIXATION (in 10% buffered formalin)

-PROCESSING (IN ALCOHOL)

-EMBEDDING (in paraffin)

-SECTIONING

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3
Q

what are the pro’s and con’s of tissue biopsy?

A

Pros: tissue RETAINS ARCHITECTURE (location in layering of the tissue is maintained)

-Allows for the use of INK to mark margins of specific area (mass)

Cons: total possible magnification is only up to 1,000X

  • ARTIFACTS OF TISSUE PROCESSING
    -when tissue is ran through ALCOHOL it shrinks (causing it to fall apart- artificial separation)
    -fats will look EMPTY because ETHANOL extracts the LIPIDS
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4
Q

What is the purpose of fixation?

A

-IMMOBILIZES components to maintain structural relationships
-prevents AUTOLYSIS
-FIRMS up structure to allow sectioning

*the faster fixation is used, the better the tissue will be PRESERVED

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5
Q

What factors affect fixation?

A

1.TIME sample is left in the fixative (small tissue 24hrs, and big tissue up to 7 days)
2. TIME UNTIL tissue is placed in fixative
3. ratio of formalin to tissue VOLUME (10:1)
4. Tissue THICKNESS (keep it thin)
-take pics and send it cut up in small pieces or BREAD loaf (cutting small 1cm slicesin big sample to allow formalin to penetrate deeper and faster)
5. tissue TYPE (blood, fat, skin, and dense connective tissue take LONGER to take up formalin)

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6
Q

UNDER fixation is a problem , as it causes cellular information to be lost

A
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7
Q

Why is decalcification required for mineralized tissue (bone and teeth)? and how is this done?

A

-calcium deposits will not section well (causes blade to become dull and chip)
-tissue is placed in ACID (FORMIC ACID, NITRIC ACID, HCL) AFTER fixation (30 mins to 7 days), AND before stain

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8
Q

what are the effects of decalcification?

A
  • damage morphology and staining properties of the tissue (proteins REMAIN but calcium is REMOVED)
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9
Q

what are the histologic stains that are most commonly used?

A

-HETAMOXYLIN: BASIC- stains negatively charged (acidic) components BLUE
-Methylene (blue)
- h&e: light purple

-EOSIN: ACIDIC- stains positively charged (basic) components PINK

-ROMANOVSKY: alcohol -> eosin stain -> difquick / wright stain NOT hematoxylin

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10
Q

What is basophilia and acidophilia?

A

-Basophilia: BLUE stain from hematoxylin
-Acidophilia: PINK stain resulting from eosin

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11
Q

Light microscopy (when is it used, what is the objective lens magnification, and ocular lens magnification)?

A

-used to evaluate ROUTINE samples
-OBJECTIVE LENSE magnification can be 2X,4X,10X, 40X, 100X
-OCULAR lense magnication multiples magnification by multiping time 10x

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12
Q

What is the different between magnification and resolution?

A

magnification is making the image LARGER, and resolution is when the clear distinction between to points on an image can be distinguished

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13
Q

What factors affect resolution?

A
  • WAVELENGTH or light (SHORTER wavelengths are better)
    -Numerical Aperture of lense (bigger lense- allows for greater collection of scattered light)
    -REFRACTIVE index of mounting media (high refractive index (oil) allows for a greater resolution)
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14
Q

when are cover class and immersion oil needed in light microscopy?

A

-cover glass are needed for samples at or greater 40X magnification (prevents blurriness)
-Immersion oil is needed starting at 100X magnification (collects more scattered light)

both improve RESOLUTION

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15
Q

How does the plane of section affect the appearance of the tissue?

A

the way that it is trimmed, placed in the cassette, and then cut, allows for different views to be observed
muscle cell : nuclei in sleeve can be seen as round, whereas on the cuff they appear cigar-shaped.

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16
Q

What is histochemistry and when is it used?

A

-it is used to highlight generic tissue that is NOT easily identified by H& E stain
-does NOT HIGHLIGHT specific Proteins
-some examples are fat, iron, copper, calcium, carbohydrates, collagen, and components go the cell walls

*** staining of cells and other tissue elements based on the chemical differences of their parts.

17
Q

what are the histochemistry pros and cons?

A

pros: picks up whatever H&E stain does not
cons: does not pick up specific proteins

18
Q

What is cytology and when is it used?

A

-common technique
-cells are aspirated through a syringe (FNA), squirt onto a slide, squashed with a second slide, AIR-DRIED, and STAINED

19
Q

what are the pros and cons of cytology?

A

pros: NONINVASIVE
-FAST RESULTS
-improved CELULLAR DETAILS
-easier to FIND ORGANISMS

-cons: ONLY picks up CELLS
-limited information (less specific diagnosis, BUT infectious agents can be seen)
-NO ARCHITECTURE

20
Q

what and when are frozen sections used?

A

-instead of fixation, embedding , etc, it is RAPIDLY FREEZE DRIED WITH NITROGEN, then stained
- it is used to get information rapidly during operation

21
Q

what are the pros and cons of frozen sections?

A

pros: rapid results

cons:
- POOR morphology (artifacts)

22
Q

What is transmission electron microscopy?

A

-sample is EMBEDED IN HARD PLASTIC
-samples are supper tiny
-samples are STAINED with HEAVY METALS
-electrons that interact with heavy metals bounce back (producing dark areas), and electrons that pass through unstained areas create the light areas

23
Q

What are pros and cons about TEM?

A

-pros: Allows for really high magnification and resolution (due to really short wave length)
-magnification of up to 10,000,000X
-cellular organelles and macromolecules (as small as 1nm) can be visualized

-cons: it is not in colo.

24
Q

What structures can you see in light microscopy?

A

-CYTOPLASM
-NUCLEUS
-NUCLEOLUS
-CELL BORDERS

25
Q

What is scanning electron microscopy and when is it used?

A
  • tissue is SPUTTER COATED DIRECTLY IN METAL (such as gold) to view the SURFACE of the tissue
    -3D IMAGE IS FORMED

OSSF (9 decks)

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