Enzyme histochemistry.pdf

Attuluri Vamsi Kumar, Assistant professor, Department of MLT, UIAHS, Chandigarh University.

1. Department of Medical Laboratory Technology, UIAHS.
Course Code: 21MSH-716 Course Name: Special Histological techniques including Immune Histochemistry and molecular pathology
Enzyme Histochemistry
Name of the Faculty: Mr. Attuluri. Vamsi Kumar E13404 Program Name: M.Sc. MLT

2. Program Structure MSc MLT 2021
Semester-I
Course
Code
Course Name LTPCCH
per
week
Course
Category
Theory
Assessment
Practical
Assessment
Total
IAMTAETACAMTAETA
21MSH-
611
Cellular
Histopathology
3034.5
6
PC
202060402040200
21MSH-
612
Applied Clinical
Biochemistry
3034.5
6
PC
202060402040200
21MSH-
613
Diagnostic
instrumentation
3034.5
6
PC
202060402040200
21MSH-
614
Clinical
Immunopatholog
y
3034.5
6
PC
202060402040200
Total Credits 1824
Cumulative Credits 1824

3. Program Structure MSc MLT 2021
Semester-II
Course
Code
Course Name LTPCCH
per
week
Course
Category
Theory AssessmentPractical
Assessment
Total
IAMTAETACAMTAETA
21MSH-651
Diagnostic
Enzymology
3034.5
6
PC
202060402040200
21MSH-652
Clinical
Immunology
3034.5
6
PC
202060402040200
21MSH-653
Molecular biology
&Genetics
3034.5
6
PC
202060402040200
21MST-654
Advance
Hematology
3034.5
6
PC
202060402040200
21MST-659
Domain Aptitude
3002
2
PC
2020600 0 0 100
Total Credits 2026
Cumulative Credits 2026

4. Program Structure MSc MLT 2021
Semester-III
Course CodeCourse Name LTPC CH
per
week
Course
Categor
y
Theory AssessmentPractical
Assessment
Total
IA MTAETACA MTAETA
21MSA-701
Blood Banking
3003
3
SE
2020 600 0 0 100
21MSA-702
Hematology
3034.5
6
SE
2020 6020 20 60200
21MSB-701
Basic and Advanced
Histological Techniques3034.5
6
SE
2020 6020 20 60
21MSH-716Special Histological
Techniques Including
Immune Histochemistry
and molecular
pathology
3033
3
SE
2020 600 0 0 100
21MSH-701
Virology and Myology
3034.5
6
SE
2020 6020 20 60

5. 21MSH-702
Medical Parasitology
3034.5
6
SE
2020 60 2020 60 200
21MSH-701
Advance Clinical Bio-
Chemistry 3034.5
6
SE
2020 60 2020 60 200
21MSH-702
Advance Techniques Used In
Clinical Bio-Chemistry 3034.5
6
SE
2020 60 2020 60 200
21MST-703
Research Methodology and
Biostatics
30 3
3
PC
2020 60 0 0 0 100
21MSR-704
Dissertation-I
0010
10
PC
00 0 4020 40 100
21MSY-710 Good Clinical Lab Practices
100 1
1
MNG
2020 60 0 0 0 100
21MSY-711 Quality Management
System
100 1
1MNG 2020 60 0 0 0 100
21MSY-712 Intricacies of NABL
100 1
1MNG 2020 60 0 0 0 100
Total Credits 21.525
Cumulative Credits 21.525
Program Structure MSc MLT 2021

6. Course Objectives
This course aims to learn the basic concepts and to get high standard knowledge of histo
related to skin, respiratory, digestive, excretory, and reproductive organ systems
Students will learn about the histopathologies of various organs of human body
This course aims to learn the basic concepts and to get high standard knowledge of histo
related to skin, respiratory, digestive, excretory, and reproductive organ systems

7. Course outcomes
CO1 Attheendofthecoursethecandidateshouldbeabletodefinethestructuralfeaturesof
respiratory,digestive,excretory,andreproductiveorgansystems.
CO2 Todevelopanunderstandingoftheanatomicalpathologiesofvariousorgansofhumanbody
CO3 The candidate should be capable of understanding cytology and histopathology of various organ
systems.Student should be capable of providing optimal pathology services in any medical set up
and develop research aptitude at PG level in the subject.
CO4 Attheendofthecoursethecandidateshouldbeabletodefinethestructuralfeaturesof
respiratory,digestive,excretory,andreproductiveorgansystems.

8. Syllabus overview
Unit.1 SpecialTechniquesForHistochemicalStudies,Principles&Advanced
SpecialStainingTechniques&enzymeHistochemistry
ContactHours:15
Chapter1
Freezedrying&vaporfixation(Specialtechniques),
Principlesoffollowing:hemoglobin,Argentaffincellgranules,Axons,Purkinjecells,Adernalmedulla,Peneth
Cells,Astrocytes,Glialfibres,Oligodendrocytes,Neurofibrills,Nerveendingandmicroglia
Chapter2
EnzymeHistochemistry:Preparationofpost–fixedandprefixedcryostatsections,Demostrationoffollowing
enzymes:
i)Phosphatase–AcidPhase,alkalinephase&ATP
ii)Dehydrogenases
iii)Oxidases
iv)Esterases
v)Tyrosinase
Experiment1
DemonstrationofdifferentfixativesusedinHisto
Experiment2
DemonstrationofdifferentMicrotomeusedinHisto
Experiment3
Todemonstratetheactivityofenzym
Experiment4
DemonstrationoffollowingenzymesactivityinaTi

9. Syllabus overview
Unit-2 Immunohistochemistry ContactHours:15
Chapter1
i)Principleofdirect,indirectandotherspecialtechniques
ii)FICandFITCtechniques
i)Techniquesofunmaskingofantigensites.
Chapter2
i)Enhancement and amplification of binding sites
ii)Double or multiple labeling techniques
iii)Preparation of conjugates.
Chapter3
i)Blockingofendogenousenzymeandbackground
ii)Methodsfoefrozenandparaffinsections
iii)Controls,internal&externalqualityassessment.
Experiment5
DemonstrationofLaboratorymethodthatusesantib
Experiment6
DemonstratetheFICandFITCtechniques
Experiment7
DemonstrationofthetechniqueusedtoseparateDN

10. Syllabus overview
Unit-3 MolecularPathology ContactHours:15
Chapter1
Molecular Pathology:
In Situ-hybrdisation: Principle, techniques and their application, Probes and Application
in fixed tissue sample.
Chapter2
PolymeraseChainReaction:PrincipleandtypeswiththeirutilizationofPolymeraseChain
Reactions
Electrophoresis:Principle,typesandtheirapplications.
Chapter3
Image Analysis: Overview, technique, specimen preparation and analysis.
Flow Cytometry: Principle, Technique, Specimen collection and preparation and their
application
Experiment8
Demonstration of technique for rapidly producing
Experiment9
Demonstration about the technique that is used for
Experiment10
Demonstrate the Flow cytometer technique used to

11. NOTES
Basics about enzymes

15. Properties of Enzymes & Factors affecting enzyme activity

19. Notes
Enzyme Histochemistry

20. Introduction

21. Significance of histochemistry

24. General Principles

25. Factors influencing the reaction

26. Factors influencing in enzyme histochemistry

27. Factors influencing in enzyme histochemistry

28. Factors influencing in enzyme histochemistry

29. Controls & Fixation

30. Fixation

31. Sections preparation

32. Principles of Enzyme histochemistry & Types of
histochemical reactions

33. Types of histochemical reactions

34. Principles of Enzyme histochemistry & Types of
histochemical reactions

35. Types of histochemical reactions

36. Principles of Enzyme histochemistry & Types of
histochemical reactions

37. Types of histochemical reactions

38. Principles of Enzyme histochemistry & Types of
histochemical reactions

39. Types of histochemical reactions

40. Demonstration of Alkaline Phosphatase

41. Clinical Significance of Alkaline Phosphatase (ALP)

42. Clinical Significance of Alkaline Phosphatase
(ALP)

43. 6.2.1Alkalinephosphatase(AKP)
AKPisahydrolyticenzyme.Ithydrolysisvariousphosphateestersofalcoholandphenol(suchassodium
β-glycerophosphateandphosphoricacidnaphthaleneester)inalkalinecondition(pH9.0–9.6).The
hydrolyticactivitycouldbeactivatedbyMg2+,Mn2+,Zn2+andCo2+andeasilyinhibitedbycysteine,cyanide
andarsenicacidsalt.Inmosttissues,theAKPcouldbeselectivelyinhibitedbytetramisole.
Demonstration of Alkaline Phosphatase (ALP)

44. Demonstration of Alkaline Phosphatase (ALP)
AKP
AKP
AKP
AKP
AKP
AKP
AKP
AKP
AKP
AKP
Alpha
Napthol
Phosphate
H20 –
pH 9.2
Napth
onol
Phosphate
Diazonium
Salt
Azopigment

46. PROCEDURE
1. Fresh tissue is fixed with 10% neutral formaldehyde for 12–24 hours or with 95%
acetone-liquid alcohol for 4–12 hours (change the fixative for two times, room
temperature)
2. Dehydrate the sections with pure alcohol, clear with xylene, embed with paraffin
and then section.
3. Dewax and rinse the sections with water for three times.

47. Syllabus overview
Unit.1 SpecialTechniquesForHistochemicalStudies,Principles&Advanced
SpecialStainingTechniques&enzymeHistochemistry
ContactHours:15
Chapter1
Freezedrying&vaporfixation(Specialtechniques),
Principlesoffollowing:hemoglobin,Argentaffincellgranules,Axons,Purkinjecells,Adernalmedulla,Peneth
Cells,Astrocytes,Glialfibres,Oligodendrocytes,Neurofibrills,Nerveendingandmicroglia
Chapter2
EnzymeHistochemistry:Preparationofpost–fixedandprefixedcryostatsections,Demostrationoffollowing
enzymes:
i)Phosphatase–AcidPhase,alkalinephase&ATP
ii)Dehydrogenases
iii)Oxidases
iv)Esterases
v)Tyrosinase
Experiment1
DemonstrationofdifferentfixativesusedinHisto
Experiment2
DemonstrationofdifferentMicrotomeusedinHisto
Experiment3
Todemonstratetheactivityofenzym
Experiment4
DemonstrationoffollowingenzymesactivityinaTi

51. Syllabus overview
Unit.1 SpecialTechniquesForHistochemicalStudies,Principles&Advanced
SpecialStainingTechniques&enzymeHistochemistry
ContactHours:15
Chapter1
Freezedrying&vaporfixation(Specialtechniques),
Principlesoffollowing:hemoglobin,Argentaffincellgranules,Axons,Purkinjecells,Adernalmedulla,Peneth
Cells,Astrocytes,Glialfibres,Oligodendrocytes,Neurofibrills,Nerveendingandmicroglia
Chapter2
EnzymeHistochemistry:Preparationofpost–fixedandprefixedcryostatsections,Demostrationoffollowing
enzymes:
i)Phosphatase–AcidPhase,alkalinephase&ATP
ii)Dehydrogenases
iii)Oxidases
iv)Esterases
v)Tyrosinase
Experiment1
DemonstrationofdifferentfixativesusedinHisto
Experiment2
DemonstrationofdifferentMicrotomeusedinHisto
Experiment3
Todemonstratetheactivityofenzym
Experiment4
DemonstrationoffollowingenzymesactivityinaTi

52. Histology of nervous system

58. FUNCTIONAL UNIT OF NERVOUS SYSTEM
IS

80. Reference
1.https://pt.slideshare.net/HusseinAltameemi2/cardiac-biomarkers-i
2.https://www.pdfdrive.com/macaca-mulatta-enzyme-
histochemistry-of-the-nervous-system-d165642716.html
3.https://www.pdfdrive.com/a-short-history-of-histochemistry-in-
denmark-e60339299.html
4.https://www.pdfdrive.com/enzyme-histochemistry-a-laboratory-
manual-d183701064.html
5.https://www.slideshare.net/KrishnenduSinha1/histochemistry-
235325874