What is the principle of Phase contrast Microscopy?

What equipment in involved in phase contract microscopes?

1. Phase condensor with annular ring

2. objective with phase shifting element.

the light rays combined producing varying degrees of contrast

enters clear ring and phase shift occurs

-Phase contrast

-Diluent: 1% ammonium oxalate

-Lyses RBC and platelets form pseudopods

-Bright field

-fix and preserve platelets and RBC

-Diluent: citrate-formaldehyfe buffer

-brillant cresyl-stain

How are platelets counted?

-using low power

-40x on large center square

-count platelets in all 25 small squares in the large center square

*each small sq. measure 0.2x0.2mm all 25 small sq is 1 large (1x1mm)

What is the normal range of platelets?

150-400 x 10^9/L

What are the sources of error in platelet counts? 

1.Light adjustment (must raise condensor)

2.Misinterpertation of bacteria and debris as platelets ( bacteria and debris are more refratile)

3. Clumping of platelets (speciment must be recollected)

4. General hemocytomer errors (overloading chamber, counting wrong borders, etc.)

Where does a platelet come from?

bonemarrow a peice of megakaryocyte

what is the average size of platelets?

2-4 um

What is the average lifespan of platelets?

7.5 days

Whate are the two major functions of platelets?

-Hemostasis 

-Maintains normal endothelial integrity

Name the events in Hemostasis

1. Vasoconstriction

2.Platelet Adhesion

3. Platelet aggregation 

4.Fibrin (Platelet plug formation)

5.Fibrin stabilization

What is vasoconstriction?

What is platelet adhesion?

-injury exposes collagen in vessel

-platelets stick to collagen (thrombus forms)

What is platelet aggregation?

What is fibrin platlet plug formation

coagularion factors interact on platelet surface to produce fibrin forming plug

What is fibrin stabalization 

Factor XIII?

What is clot retraction?

Platelet-fibrin clot contracts (requires lots of ATP)

Thrombothenin= contractile protien in platelets

In Vitro: clot pulls away from tube leaving clear serum 

-requires a few hours

What is involed in vascular repair?

platelet dervied growth factor: promotes smooth muscle growth for vessel  repair.

B-thromboglobin: chemotactic agent for fibroblasts to help with vessel repair

*decreased platelet production

--congenital drugs, chemo, marrow displacement

*Increased consumption

-increased hemostasis

*increased sequestration 

-hypersplenism

*increased destruction

-idiopathic thrombocytopenic purpura

What is thrombocytosis?

Primary

-associated with myeloproliferative disorders (abnoral production of one of more elements from bone marrow)

Secondary

-iron deficiency anemia, hemorrhage, hemolysis, splenectomy, inflammation

decrease in function associated with abnormal bleeding

(thromboasthenia (glanzmanns) failure of platelets to aggregate in presence of ADP, invivo -defect in formation of hemostatic plugs and serious bleeding episodes.)

Secondary functional platelet defects

decrease in function due to drugs, aspirin, & uremia

Platelet count requires which anticoagulant

EDTA

1:10 to 1:40

7-25

*multply # by 20,000 to approximate manual or automated platelet count