Download MBBS 2nd Year Clinical Pathology Notes Notes

CLINICAL PATHOLOGY

PERIPHERAL BLOOD SMEAR

Importance of PBS: PBS (Peripheral blood film) is a very valuable & frequently asked question in hematology. It provides the following information:

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1. Red cell morphology:
   • Morphological features of RBCs like size, shape, colour, the presence of inclusions - important for diagnosis of Anaemias & other hematological disorders.

Variation in size of RBC:

Colour of RBC:

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2. WBC disorders & DLC:
   • Quantitative & qualitative changes in WBC suggests helps in diagnosis of hematological & non-hematological disorders.
3. Platelet count & morphology:
   • Useful in the diagnosis of bleeding disorders.
4. Cross check the CBC parameters: helps in cross checking the CBC parameters, derived from automated cell counters.

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5. CBC/Hemogram
   1. Hb
   2. TLC
   3. TEC
   4. TPL
   5. DLC

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   6. Hematocrit/PCV
   7. Reticulocyte count

PBS

• RBC morphology

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• Platelet
• Hemoparasites
• Any atypical / abnormal cell

6. Detection of blood parasites (Hemoparasites)

Hemoparasites include:

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1. Malarial parasite i.e. Plasmodium
2. Microfilaria
3. Trypanosomes
4. Leishmania donovani
5. Kala-Azar

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RETICULOCYTE COUNT:

Reticulocyte - immature, non nucleated RBC released from B.M. slightly larger than mature RBC.

Continue to synthesize Hb, after loss of nucleus.

Normal Reticulocyte count: expressed in terms of % of total RBC

N - 0.5-2.5%

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Importance of Reticulocyte counting:

Reticulocyte count seen

Following increase in:

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1. IDA
2. FA > def anemia
3. Vit B12

* highest count is seen on 6/7th day & indicates BM. response to hemorrhage.

3. Primary & 2° polycythemia

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1. Anemia due to organ failure - Renal failure & hepatic disorders
2. Pure Red cell Aplasia
3. Fanconi Anemia
4. Myelofibrosis

* In short, Reticulocyte count is a significant test during investigations of various hematological disorders.

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1. It can almost accurately reflect hematopoietic activity.
2. It can help in diagnosis & differential diagnosis of anemias & other diseases.
3. It can assess the therapeutic response of different anemias.

Reticulocytosis - can be a cause of peripheral macrocytosis & normoblastic hematopoiesis; as commonly seen in hemorrhagic disorders.

HEMATOCRIT (HCT)/ Packed cell vol (PCV)/ Erythrocyte volume fraction.

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Definition: is the ratio of vol of RBC : whole blood.

• It indicates the relative volume of RBC & plasma.
• In anaemia ? RBC ? ? hematocrit

Method of Estimation of Hct/PCV:

1. Macro method using Wintrobe's tube.

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2. Micro method using capillary tube
3. Automated analyzer.

Wintrobe's method:

Wintrobe's tube is a special thick walled glass tube -

l - 11 cm

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d (interval) - (2.5mm)

capacity - (1ml)

Calculated (0 ? 100) - ESR

upside down (100 ? 0) - PCV estimate

Anticoagulants descending order used

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• EDTA
• dried heparin
• double oxalate

Principle: Anticoagulated whole blood is centrifuged at std. speed. RBC > WBCs, platelets, plasma settle down at the bottom & hematocrit indicates the hematocrit vol of RBC.

Different layers:

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Upper layer: clear/ viscous fluid straw coloured plasma.

Middle layer: thin Red grey layer. (platelets & WBC.

Plasma

RBC

Lower layer: RBC (packed)

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Normal Range of PCV/Hct: expressed in % or decimal fraction (0.95).

Useful for estimating absolute values like MCV

Adult ? - 38-47%

Adult ? - 36-46%

Infant - 44-45-10% (cord blood)

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PCV.

1. Polycythemia vera rubra
2. Secondary polycythemia

Bleeding Time (BT)

(2-9 min.)

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Normal Range:

Definition: time interval b/w oozing of blood from cut/injury till stoppage of bleeding.

Method:

1. Duke's method (absolute)
2. Ivy's method

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3. Template method-method of charm

Template Method for B.T.:-

Template: disposable blade, fitted on to a holder.

made up of plastic, used for test

recommended as routine pre-operative screening test.

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BT - usually ? prolonged in pt. coagulation factor deficiency.

Prolonged B.T. is found in:

1. Platelet disorder. PIC(NPC < 50k /mm³)
   • Quantitative: Thrombocytopenia
   • Qualitative: vW disease.

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Coagulation / Clotting Time (CT)

(4-11 min)

1. time interval from b/w oozing of blood from cut / injury till formation of clot.
2. Lee-White Method (absolute now.)

Disadvantages:

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1. Not sensitive: one of the oldest tests & ? used now
   • fails to detect mild/moderate coagulant defects.
   • absolute now ? recommended as screening Test.

PTT- more sensitive test for assessment of coagulation cascade.

4. Misleading Normal values may be obtained in mild ? moderate Hemophilia A & B.
   • ? exclude major factor deficiency

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Bombay Blood Group

special type of blood group, where there is no H-substance, expression over RBC.

* H is the basic precursor Ag for native A & B Ag. So, it is one of the Ag of ABO blood grp. system

Basic precursor substance.

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A gene H-gene H-substance

Transformed

A Ag B gene A, B Ag

B Ag

H

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Genes (the 'Ag' expressed over RBCs Blood Grp

Precursor substa

A A, H A

H B B, H B

A, B A, B, H AB

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H H O

* Usually, RBCs from all persons contain H-Ag on their surface, so, in the presence / -nee of A & B gene H Ag will transform into, A, B, AB & H Ag (no change)

Depending on these types of ABO blood grps are categorized.

In case of Bombay blood group no H gene not-rules. I'H'Ag

So even in the presence of A/B gene, no A/B Ag will be expressed over RBC surface".

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RBC will ? express any form of ABO Ag (A, Both).

During forward / classical blood grouping, there will be no clumping of test blood & anti A/B sera. So, the result will mimic O blood group.

Problem arises & preformed serum Abs.

In ABO blood grp. system, plasma will contain preformed Abs against Ent Ag.

So. Blood grp A Anti-B capable of Agglutination

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B Anti-A

AB no Abs

O Anti A, B.

as all blood grp contain H Ag - So, no Anti-H.

Agglutination

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But Bombay blood grp will have Anti-H Abs in their serum. So, they show transfusion reaction will all other blood grps of ABD system i.e., A, B, AB, O.

This agglutinating serum Abs-demonstrated by reverse blood grouping.

Importance of Bombay Blood grp:

1. Bombay blood grp pt can accept blood only from Bombay blood grp donor.

2. A register enrolling Bombay blood grp carrier-must be maintained state & national level, for collection of blood, when necessary

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ABO Blood Grouping Technique

1. Type Angulation of

Forward (cell typing) Reverse (serum typing)

Ag on RBC-detected Abs in serum/

plasma-detected

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Reverse blood Grouping- Usually done by tube method

Procedure:

Patient serum

+

Reagent (RBC) of known blood grp.

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(supplied / prepared)

Incubate- 5min at room temp.

centrifugation

sedimentation of RBC at the bottom

of test tube

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Resuspension of sedimented RBCs

by gentle tapping

clumps of RBC Uniform suspension of

suspended in clear fluid RBC

+ve Test -ve Test

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anti RBC Abs test (Serum Abs test)

seperate tubes for +ve & -ve controls are used.

By reverse blood grouping serum Abs against RBC surface Ag can be detected.

routine forward blood grouping can only detect Ag. the on the surface of RBC, using known Anti A/B Abs.

this method can miss out anti RBC Abs in serum of ABO blood grp. that in serum, which may lead to transfusion hazards. I'd as 'O' blood grp ( no ABO Ag on the surface of RBCs).

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But,

it Bombay blood grp transfused to 'O' blood grp

Hemolysis

(: Anti 'H' Abs mt in recipient's serum).

Reverse blood grouping - demonstrates the three of serum Abs (anti H Abs) against all types of RBC of ABD system

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transfusion hazards can be avoided.

Other advantages of reverse blood grouping are:-

1. Easier i'd of weakly reactive Ag (like a 1).
2. A method of double checking, to rectify any flaw of forward grouping, as mentioned above
3. to detect serum and RBC Abs, which may be missed during usual grouping

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4. v. imp in cases, where there will be pathological change in antigenicity of RBC i.e. colonic carcinoma

Transfusion Related disease can be avoided

A lot of infective diseases can be transmitted during blood transfusions like:-

1. Bacterial: Syphilis.
2. Viral - Hepatitis-B, C, D & HIV-I & II

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3. Parasitic - Malaria

For transmission to occur, the donor must be a sufferer/carrier of the disease

Nowadays, adequate tests are available for the detection of infection in donor's blood & unless seen to the devoid of all contaminants? used for transfusion.

Problem arises when blood is collected during incubation period of infection.

usual screening term may ? +ve.

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Ex- during early phase of HIV infection

ELISA may be -ve

ELISA +ve case.

further examined for the the of P-24 antigenemia

will be +ve even during

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earliest phase of infection

In this way, almost all infected carriers can be detected, irrespective of the duration of disease.

Ave blood samples healthy, disease free samples are destroyed preserved for future blood transfusion.

So, safe blood banking procedure, transfusion related diseases can be avoided.

Blood component therapy is beneficial than whole blood transfusion

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After collection of blood donor, blood is divided into different components by centrifugation.

whole blood.

1st centrifugation

Packed RBC/ Platelet Rich

Red cell concentrate Plasma.

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2nd centrifugation

Platelet Platelet poor plasma

concentrate Fresh plasma

FFP

These components are stored in appropriate conditions & used for transfusion to suitable recipient a/c need.

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Cryoprecipitate

• include
• plasma pr
• fibrinogen
• Factor VIII

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• VWF

The main advantage of component therapy, over whole blood transfusion are:

1. A single unit of blood can be used for benefit of >1 pt. So, it is cost effective. Ex. RBC can be transfused to an anemic pt. & plasma to burn pt.
2. It can ensure that only the required component are transfused. Ex - During Ht of anaemia, transfusion of whole blood vot overload heart failure.

Packed RBC transfusion is free from all these hazards.

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3. Component separation helps to maintain the potency of individual component for longer period. Ex-whole wood no platelet.

after 48 hrs

• platelet concentrate can be used for preserved 5-6 days
• Cryoppt. & FFP can be used for much longer period

Component separation - tse the wastage of donated blood. chance at least one/2 component & single unit chance of whole blood utilisation

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so, component separation should be encouraged in blood banking.

Fresh Frozen Plasma (FFP)

prepared by freezing the plasma.

contains plasma proteins & all coagulation factors

that, include Albumin

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Protein C

Protein S

Santh thrombin

VWF

It is indicated in replacement of coagulation factors, in acquired coagulation factor deficiency

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Indications for FFP:

1. Pt on anticoagulant drug therapy (ex. Coumarin)
2. vit k deficiency
3. Antithrombin Deficiency
4. coagulopathy of liver disease

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5. Microangiopathic hemolytic Anemia, including TTP, hemolytic uremic syndrome, HELLP syndrome
6. DIC.

TRANSFUSION REACTIONS.

Blood transfusion is useful & lifesaving when performed a caution & clear indication.

St. (~2-4% of cases), unfavourable complications can occur, in spite of precision & preventive measures,

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which are k/a transfusion reactions.

Complication of blood transfusion/

Transfusion reactions.

Infectious complications Non-infectious complications

some disease transmitted by transfusion are.

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1. Hepatitis (HBV, HCV, HDV).
2. HIV (AIDS)
3. Cytomegalovirus
4. Malaria.
5. Syphilis.

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Immediate Rxn Delayed Rxn

Immunological Non Immunological Non immunological immunological

Hemolytic circulatory Alloimmunz De-overload

transfusion rxn. overload - ation -ad transfusion

febrile non- Air hemolytic Thema-

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hemolytic rxn. embolism. transfusion - siderosis

Allergic rxn rxn (mostly thrombos

Anaphylactic asymptomatic), -phlebitis.

TRALI Transfusion-

Transfusion related associated graft-versus

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acute lung injury. host disease.

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