Histo Intro (Part 2)

Front 1

It acts as a permanent record of tissues, organs, and specimens.

Back 1

Slides

Front 2

What are the info needed to provide in a slide?

Back 2

1. Name of the organism then w.m. or e

2. Part of the organism used

3. Type of preparation

4. Date

5. Signed

Front 3

The process of mechanical cutting of plant/animal materials are done by the aid of microtomes

Back 3

Microtomy

Front 4

Four basic groups of Microtomy

Back 4

1. Rocking microtome

2. Rotary microtome

3. Sledge microtome

4. Freezing microtome

Front 5

generally best for cutting sections thicker than six microns.

Back 5

Rocking microtome

Front 6

This microtome is not suitable for embryological specimen specially if the juxtaposition of the cells in critical or when doing serial section

Back 6

Rocking microtome

Front 7

More expensive than rocking microtome

Back 7

Rotary microtomes

Front 8

enable sections of 5 microns thickness to be cut.

Back 8

Rotary microtomes

Front 9

May weigh as much as fifty kilograms, and consists of a heavy base into which aremachined flat tracks on which the chuck sledge slides.

Back 9

Sledge Microtome

Front 10

used to cut sections down to two or three microns.

Back 10

Sledge Microtome

Front 11

This device usually has a wedge blade

Back 11

Freezing microtome

Front 12

Such microtomes are sometimes mounted in chilled cabinets to make working with themeasier

Back 12

Freezing microtome

Front 13

now possible to cut frozen sections using freezer aerosols instead of CO2 andfreezing chucks are made which are electrically cooled by means of Peltiercells.

Back 13

Freezing microtome

Front 14

three basic cross sectional shapes of knives

Back 14

1. wedge

2. plano-concave

3. double concave

Front 15

Knives that is generally,though not exclusively used for cutting frozen sections

Back 15

Plane wedge knife

Front 16

Knives are often used for sectioning soft materials such as celloidin embedded tissues.

Back 16

plano-concave

Front 17

razors are generally plano-convex.

Back 17

Botanical sectioning

Front 18

best knives for the cutting of paraffin blocks

Back 18

double concave type

Front 19

Combine histological methods with chemical or biochemical methods, make some compositions of tissue or cell become insoluble, colored or electron-dense, to show those chemical compositions of tissue or cell in situ, such compositions includes protein, amino acid, nucleic acid, lipid and enzymes.

Back 19

General Histochemistry

Front 20

¨branch of science concerned with the qualitative and quantitative assessment of chemical compounds in a cell/tissue using stains/dyes and microscopy.¨

Back 20

Histochemistry

Front 21

marriage between bio/chemistry and cyto/histology

Back 21

Histochemistry

Front 22

¨He stained starch in plant tissues blue with potassium iodide solution under the light microscope and demonstrated its localization microscopically

Back 22

Raspail,1825

Front 23

he published an essay on microscopic chemistry for the first time

Back 23

Raspail,1825

Front 24

¨first to introduce cell fractionation to analyze nucleic acids in nuclei of leukocytes. ¨

Back 24

Miescher(1874)

Front 25

It is used to analyze nucleic acids in nuclei of leukocytes.

Back 25

Cell fractionation

Front 26

¨ microscopic chemistry or chemical microscopy are meant to observe chemical reactions in situ under microscopy.

Back 26

Microchemistry

Front 27

¨frequently used to stain tissues in anatomy and pathology during early 20th century

Back 27

Aniline dyes

Front 28

published his famous book entitled “Histochemie Animale”,

Back 28

Lison(1936)

Front 29

¨He proclaimed Histochemistry to be the new science

Back 29

Lison 1936

Front 30

¨proposed that the field should be designated General Histo-cytochemistry just like we have General Histology.

Back 30

Nagata(1995)

Front 31

One of the foremost histochemists in Nigeria

Back 31

LateProf. Caxton-Martins

Front 32

classificationof histochemical methods

Back 32

1. Chemical methods such as chemical reactions by staining

2. Physical methods such as radiations

3. Biological methods such as immunity

Front 33

¨histology,anatomy, immunology and biochemistry to detect the amount, distribution andlocalization of a specific target within a tissue.

Back 33

Immunohistochemistry (IHC)

Front 34

anumbrella term that encompasses all the techniques that are used for thedetection of molecules employing the antigen–antibody reaction

Back 34

Immunostaining

Front 35

aspecific use case of immunostaining when the antigen–antibody reaction is usedto study the status of molecules in tissue

Back 35

Immunohistochemical staining

Front 36

¨Immunohistochemistryrefers to the evaluation of target antigens in tissues whenthe detection method can be either chromogenic or fluorogenic.

Back 36

Immunohistochemistry

Front 37

mayrefer to both the evaluation of target antigens in cells and tissues andspecifically involves the detection of a fluorescent label(fluorophore)

Back 37

Immunofluorescence

Front 38

studyof the status of antigens in tissue samples

Back 38

immunohistochemistryi

Front 39

immunostainingwhen the sample of interest is cells in culture

Back 39

immunocytochemistry

Front 40

Parameters in Performing Immunohistochemistry

Back 40

1. tissue

2. target

3. epitope

4. fixation method

5. fixative

6. sample preparation methods

7. sectioning methods

8. pre-processing tissue sections

9. antigen retrieval methods

10. permeabilization

11. blocking buffer

12. detection method

13. primary antibodies

14. secondary antibodies

15. signal amplification

16. labels

17. counterstain

18. mounting

19. multiplexing

20. imaging method

21. controls

Front 41

thesmall three-dimensional surface region of the antigen to which an antibodywould specifically bind.

Back 41

epitope

Front 42

Fixativesfall into three categories:

Back 42

1. aldehyde

2. alcohols

3. acetone-based fixatives

Front 43

mostcommonly used fixative is

Back 43

4%(w/v) paraformaldehyde solution prepared in phosphate buffered saline (PBS).

Front 44

Twocommon methods of sample preparation

Back 44

1. formalin-fixed paraffin embedding (FFPE)

2. freezing.

Front 45

Antigenretrieval can be achieved either by the application of

Back 45

1. heat(heatinduced epitope retrieval: HIER)

2. enzymatic degradation (proteolytic-induced epitoperetrieval: PIER)

Front 46

surfactants used to achieve permeabilization

Back 46

1. TritonX-100

2. Tween-20

3. saponin and digitonin

Front 47

The concentration of the surfactant and the time of incubation are determinedbased on factors such as the

Back 47

1. fixative used

2. thickness of the tissue section

3. subcellular localization of the antigen of interest.

Front 48

Commonly used blocking agent

Back 48

bovine and serum albumin

Front 49

Primary antibodies may be

Back 49

polyclonal or monoclonal

Front 50

Severalsignal amplification strategies that are used includ

Back 50

1. avidin-biotincomplex (ABC)method

2. labeledstreptavidin-biotin (LSAB)method

3. tyramide signalamplification(TSA).

Front 51

Labels used to detect the target antibody are attached to the primary or secondary antibodies and may be either _ or _

Back 51

1. fluorogenic

2. chromogenic.