Laboratory investigation

1. Laboratory
Investigation
PRESENTED BY-
DR. AASTHA
SAHANI
DR. DIMPLE
CHAWLA
DR. MAHIMA JAIN

2. Content
• Need for laboratory investigations
• Definition
• Generic application
• Classification
• Laboratory investigation
a) Haematological investigation
b) Biochemistry investigation
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3. Cont.
c) Microbiological investigation
d) Immunological investigation
e) Histopathological investigation
• Common clinical scenarios
• Conclusion
• References
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4. Need for
Laboratory Investigation 4
• Evidence shows Case history
and clinical examination usually
revel most, if not then we use
investigation
• Some systemic conditions of
which patients are unaware of
are often identified.
• To confirm our clinical
impression.

5. Definition
• LABORATORY STUDIES ARE AN EXTENSION OF PHYSICAL
EXAMINATION IN WHICH TISSUE, BLOOD, URINE OR OTHER
SPECIMEN ARE OBTAINED FROM PATENTS AND SUBJECTED TO
MICROSCOPIC, BIOCHEMICAL, MICROBIOLOGICAL OR
IMMUNOLOGICAL EXAMINATION.
• INFORMATION OBTAINED FROM THESE INVESTIGATIONS HELPS IN
IDENTIFYING NATURE OF THE DISEASE.
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6. Generic Applications
• Confirming or rejecting clinical diagnosis
• Providing suitable guidelines for patient management
• Providing prognostic information of the disease under
consideration
• Detecting disease through case-finding screening methods
• Monitoring follow up therapy
• Provides info for Medico-legal consulatiations.
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7. Classification
Based on where investigation is done:
Chair side Investigation
• Acts as a precurser to laboratory
investigation.
• Eg. Toluidine staining for grading
dysplasia, Electric pulp testing for
tooth vitality.
Laboratory Investigation
• Significantly higher sensitivity
and specificity
• Eg. Glycated hemoglobin
estimation, Peripheral blood
smear.
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8. Based on specificity and sensitivity:
Screening test
• An ideal screening test is 100%
sensitive
• Useful in large sample size at
risk; typically cheaper
• Eg. Blood glucose estimation ,
Hematocrit values for anemia,
VRDL test for syphilis.
Diagnostic test
• An ideal diagnostic test is 100%
specific
• It is expensive
• Hb1ac, OGTT peripheral smear
histology.
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9. Based on hospital lab services:
• Hematology
• Microbiology
• Biochemistry
• Histopathology
• Cytopathology
• Immunology
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10. Haematology
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11. Definition
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Deals with
abnormalities of
blood cells, their
precursors and of the
hemostatic and
clotting mechanism.

12. Haematological
Investigations
• Hb
• PCV
• RBC COUNT
• TLC
• DLC
• PLATELET COUNT
• ESR
• RBC INDICES
• PROTHROMBIN TIME
• ACTIVATED PARTIAL
THROMBOPLASTIN TIME
• SERUM IRON AND TOTAL IRON
BINDING CAPACITY

13. RBC count
• Normal range- Adult male: 4-6 million cells/cu.mm
Adult female: 3-5 millions cells /cu.mm
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Polycythemia Anemia
• Abnormally high values of circulating RBCs.
• May be primary or secondary
• Abnormally low values of circulating RBCs
• Seen in abnormality of bone marrow ( primary)
or altitude related ( secondary )
• May result from chronic hemorrhage, bone
marrow failure.

14. Haematocrit
• Volume of packed erythrocytes/100 ml of blood done in a
centrifuge.
• Although test is inaccurate, it is more precise than the
erythrocyte count and is used in combination with it.
• Normal range: Adult male -40-50%
Adult female- 38-42%
• These values are increased in polycythemia and reduced in
anemia.
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15. Haemoglobin ( Hb)
• Oxygen carrying component of erythrocyte.
• Normal range: Adult male- 14-18%
Adult female- 12-16%
Low value indicates anemia and high values indicate
polycythemia.
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16. Peripheral smear
• Provides information concerning the size and shape of
RBC.
• It may allow identification of sickle cell anemia &
macrocytic,microcytic and normocytic anemia.
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17. Mean Cell Volume ( MCV)
• Ratio of Haematocrit to RBC count expressed in 𝜇𝑚3
• Describe volume of RBC.
• Range : Normal-82-90 𝜇𝑚3
Normocytic anemia- 50-80/𝜇𝑚3
Microcytic anemia- 50-80/𝜇𝑚3
Macrocytic anemia- 95-100𝜇𝑚3
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18. Mean Cell Haemoglobin ( MCH)
• Ratio of Hb to RBCs and is expressed in picograms.
• It expresses the Hb content of each cell.
• Range: Normal- 27-31pcg
Normocytic anemia- 15-25 pcg
Microcytic anemia- 15-25 pcg
Macrocytic anemia- 30-50 pcg
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19. Mean Cell Haemoglobin Concentration (
MCHC)
• Ratio of Hb to Hb concentration.
• Value expressed as percentage of volume of blood cells.
• Measures Hb concentration in grams/100 of packed
erythrocytes.
• Range
1. Normal-32-36%
2. Normocytic anemia- 32-36%
3. Microcytic anemia- 25-30%
4. Macrocytic anemia- 32-36%
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20. Erythrocyte sediment rate ( ESR)
• ESR indicates the speed with which the erythrocytes settle in
uncoagulated blood
• Values :
1. Men < 50 years- <15 mm/hr
2. Women < 50 years- <20 mm/hr
3. Men > 50 years- <20 mm/hr
4. Women > 50 years- <30 mm/hr

21. Interpretation
Raised ESR
• TB
• SABE
• ACUTE MI
• SEPTIC SHOCK
• ANEMIAS
Lowered ESR
• POLYCYTHEMIA
• SPHEROCYTOSIS
• SICLE CELL ANEMIA
• CONGESTICE HEART FAILURE
• NEW BORN INFANT
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22. White blood cell count
• White blood cells are classified as Granulocytes and
Agranulocytes.
• Normal range: 4500-11000 cells/m𝑚3
• High values may be caused by Leukemia, polycythemia or
infectious disease.
• Low values may be due to bone marrow depression, aplastic
anemia, drug reaction and viral infections.
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23. Differential Leucocyte Count (DLC)
• Obtained from peripheral blood smear.
• Values are interpreted in total percentage of WBC.
NEUTROPHILS:
• Band neutrophils are immature and seg neutrophils are mature.
• Normal band value – 2-3% while normal seg value – 50-60%
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24. Cont….
Basophills:
• Normal value- 0-1%
• High values uncommon: may indicate myeloproliferative disease.
• Low values may indicate an oncoming anaphylactic reaction.
Eosinophills:
• Normal value- 0-5%
• High values are mostly observed in allergies or parasitic infection.
• Low values are mostly observed in aplastic anemia.
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25. Lymphocyte:
• Normal value- 30-40%
• High value may indicate viral infection, lymphocyte leukemia
• Low value may indicate aplastic anemia.
Monocytes:
• Normal values 3-7%
• High values are seen in monocyte leukemia, Hodgkin’s disease,
SABE.
• Low value are mostly seen in aplastic anemia.
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26. Bleeding Time
• Measures the time for haemostatic plug formation.
• Normal bleeding time- 2-7 mins
• Any clotting factor deficiency or platelet abnormality will lead to
increase BT.
• Prolonged in:
i. Thrombocytopenia
ii. Acute leukemia
iii. Aplastic anemia
iv. Liver disease
v. Von- Willebrand’s disease.
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27. Clotting Time
• Measures the time required for formation of clot.
• Screening test for coagulation disorder
• Normal Clotting time- 4-14 mins
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28. Prothrombin Time
• Required for fibrin threads to form in citrated or oxalated
plasma.
• Normal time- 11-14 sec
• Measured against a control PT in terms of INR.
• Measures extrinsic and common pathway- Factors I,II,V,VII,X.
• Increased PT:
i. DIC
ii. Patient on warfarin therapy
iii. Vit k deficiency
iv. Liver failure.
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29. Activated partial thromboplastin time (aPTT)
• Time in seconds that’s required for a clot to form in citrated or
oxalated plasma.
• Performance indicator of both the intrinsic & common pathways.
• Typical reference range- 30-40 secs
• Increased aPTT seen in:
i. Patient on heparin therapy
ii. Von- willebrand’s disease.
iii. Haemophilia
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30. BioChemistry
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31. Definition
• Also called chemical pathology
• Deals with investigation of the metabolic abnormalities of the
body in disease state.
• Investigations are carried out by assays of various normal and
abnormal compounds found in body fluids.
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32. Serum
chemistry
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• Serum is that portion of
blood remaining after
whole blood has been
allowed to clot
• Responsible for fluid
maintenance intra and
extra cellularly
• Responsible for the
optimal osmotic gradient,
nerve and muscle
function and hydration.

33. Blood glucose estimation
• Fasting blood glucose: 70-90mg/100ml
• Random blood glucose: 110-130mg/ml
• Post prandial blood sugar: <140 mg/100ml
• High values indicate diabetes mellitus
• Low values are seen in insulin secreting tumours.
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34. Oral Glucose Tolerance Test
• Used for definitive diagnosis of diabetes mellitus and distinguish
diabetes from other cause of hypoglycemia like
hyperthyroidism.
• Should be performed on only healthy patients who are not
under any drugs which may interfere with glucose estimation.
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35. Glycated haemoglobin ( HbA1c )
• Hb becomes Glycated by keto-amine reaction between glucose
and other sugars.
• It provides a definitive value of blood sugar control for 2-3
months.
• It is considered to be a better indicator for diabetic control
compared to blood glucose level.
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36. Range
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37. Serum Calcium, Phosphorus
• Indicated on suspicion of Paget’s disease, fibrous dysplasis,
primary and secondary hyperthyroidism, osteoporosis, multiple
myeloma or osteosarcoma.
• Concentration of Ca varies inversely proportional to
concentration of P.
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38. Range
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• Normal level serum Ca – 9.2-11 mg/dl
• Normal level serum P – 3-4.5 mg/dl
• At levels less than 7mg/dl serum Ca, signs of tetany may
appear

39. Serum Alkaline Phosphatase ( ALP )
• ALP produced in small amounts in the liver but most notably on
osteoblasts.
• Range
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HIGH VALUES LOW VALUES
OBSTRUCTIVE LIVER
DISEASE
HYPOPHOSPHOTASIA
PAGET’S DISEASE HYPOTHYROIDISM
OSTEOMALACIA OSTEOPOROSIS
RICKETS APLASTIC ANEMIA
SARCOIDOSIS CML
LYMPHOMA WILSON’S DISEASE

41. This test is very useful for
diagnosing biliary obstruction.
• EVEN IN MILD
CASES THIS
ENZYME IS
ELEVATED.

42. Serum Uric Acid
• End product of purine metabolism.
• Abnormally high uric acid level seen in gout, renal failure,
leukemia, lymphoma, starvation, poisoning, cancer
chemotherapy.
• Low values are rare.
• Range:
i. Males: 2.1-7.8 mg/100ml
ii. Females: 2.0-6.4 mg/100ml
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43. Serum Creatinine
• Metabolic product of dephosphorylation of creatinine
phosphate.
• Raised in late stages of renal disease.
• Levels > 15mg/dl indicates impaired renal metabolism.
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44. Blood Urea Nitrogen
• Formed by deamination of of amino acid in the liver.
• Protein metabolism produces ammonia, a toxic substance that
is converted into urea.
• Normal values : 8-18 mg/dl.
• High values are seen in acute and chronic renal failure,
congestive heart failure and urinary tract infection.
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45. Total Protein & Albumin/Globulin ratio
• These are important in coagulation, transport a variety of
hormones, act as buffer systems and help maintaining osmotic
pressure.
• Normal ranges:
• Total protein: 6-8g/dl
• A/G ratio: 1.2-2.0
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High total protein
values
Low total protein
values
LUPUS
EYTHEMATOSUS
INADEQUATE
PROTEIN INATKE
COLLAGEN DISEASE PROTEIN
MALABSORPTION
ACUTE LIVER
DISEASE
DIARRHOEA
MULTIPLE MYELOMA ANEMIA AND BURNS

47. Serum bilirubin
• Bilirubin is the bile pigment derived from the breakdown of
Haemoglobin.
• Normal values: 0.1-1.2 mg/100ml
• Levels >3 mg/100ml may indicate jaundice.
• High values also indicate haemolytic anemia, biliary destruction,
hepatitis and Gilbert’s disease.
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48. LDH, SGOT, SGPT
These enzymes can be indicative of liver disease.
LDH
• Responsible for the
oxidation of lactic acid to
pyruvic acid.
• Normal range: 71-207
IU/L
SGOT (AST)
• Responsible for
conversion of amino acid
to keto acids
• Normal range: 0-35 IU/L
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SGPT ( ALT)
• Responsible for diagnosis
of liver function more so
that SGOT levels.
• Normal range: 0-35 IU/L

49. Blood electrolytes
• An automated analysis usually includes Sodium, Potassium,
Bicarbonates and chlorides
• Normal values :
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SODIUM 136-145 mEq/L
POTASSIUM 3.8-5.5 mEq/L
CHLORIDE 95-105 mEq/L
BICARBONATES 22-28 mEq/L

50. Saliva
chemistry
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• Secretions are
collected directly from
individual parotid,
sublingual and
submandibular glands
by one of the small
rubber cups pressed
slightly against gland
orifices.

51. Normal values of unstimulated saliva
• K-25 mEq/L
• Na- <10 mEq/L
• Cl- 15-18 mEq/L
• Increased K or Na values may indicate generic inflammation or
sialodenosis.
• In Sjogren Syndrome:
Salivary flow rate is reduced
Na,Cl conc. Is reduced.
Salivary IgA is elevated.
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52. Microbiology PIC

53. Microbiology
• Culture and sensitivity tests are used to isolate and identify
causative organisms of an infection
• May be obtained from blood and urine.
• Particularly helpful in evaluating infections related to throat,
sinuses, root canal or bone.
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54. Immunology
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55. Immunofluorescence
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• This procedure employs the
use of fluorescent labelled
antibodies to detect specific
Ag-Ab reaction of known
specificity in tissue sections.
• When tissue section labelled
in the fashion are illuminated
with ultra violet light in UV
microscope, specific labelled
tissue component can be
identified by their bright apple
green fluorescence against a
dark background.

56. Types of Immunofluorescence
Direct
• Adding of fluorescent
labelled Ab to patient
tissue
• Wash
• Visualize under
fluorescent microscope.
Indirect
• Addition of patient
serum to tissue
containing known Ag
• Wash
• Add fluorescent
labelled Ag.
• Wash
• Visulaize.
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57. Procedure
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58. ImmunoPrecepitation Assays
• Detects Ab in solution
• End point is visual flocculation of the antigen and the antibody
in the suspension.
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59. Compliment Fixation
• Based on activation/fixation of complement following binding of
complement factors to Ag-Ab immune complexes.
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60. Particle agglutination
• Relatively simple and fast.
• Capable of detecting lower concentration of antibodies.
• Designed to detect antibodies to viruses, subsequent to
vaccination.
• Utilizes Ag coated latex particles, coal particles.
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61. Enzyme Immuno Assay
• Most sensitive
• Depends on anti human IgG or IgM Ab conjugate
• Ab conjugate, if present is made to attach to enzyme which
catalyses conversion of substrate to colored product which is
then read by spectrophotometer
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62. Radio Immuno Assay
• Extremely sensitive and specific procedure.
• Measure conc. of Ag in patient sera by Ab
• A known quantity of Ag is made radioactive and is made to
compete with Ag in patient’s sera for Ab binding sites.
• The radioactivity of free Ag remaining is measured using
Gamma counter
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63. Histopathology
&
Cytopathology

64. Histopathology
• Refers to the microscopic
examination of tissue in order to
study the manifestation of the
disease.
Cytopathology
• Refers to scientific study to role
of individual cells or cells type in
disease.
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65. Tissue Biopsy
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• A biopsy is a controlled
and deliberate removal of
tissue from a living
organism for the purpose
of microscopic
examination.
• Relatively simple
procedure producing little
discomfort when
compared to exodontia or
periodontal surgery.

66. Uses
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• Diagnosis
• Grading of tumors
• Metastaic lesion
• Recurrence
• Management assessment

67. Tissue Biopsy Types
Excisional
biopsy Incisional biopsy
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Punch biopsy

68. Exfoliative
cytology
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• The surface of the
lesion is either
wiped with a
sponge material
or scraped to
make a smear.

69. Interpretation of Exfoliative Cytology
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70. Fine Needle
Aspiration Cytology
• MICROSCOPIC
EXAMINATION
OF AN ASPIRATE
OBTAINED BY
INSERTING A
FINE NEEDLE
INTO A LESION

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