BIOT 309: BLOOD SMEAR PRACTICAL Feb., 2013. Microscopic Views Bird Blood Cat Blood Dog Blood Fish Blood Frog Blood Snake Blood Human Blood Horse Blood.

BIOT 309: BLOOD SMEAR PRACTICAL

Feb., 2013

Microscopic Views

Bird Blood

Cat Blood

Dog Blood

Fish Blood

Frog Blood

Snake BloodHuman Blood

Horse Blood

Maslak, P. ASH Image Bank 2008;2008:8-00067. Copyright ©2008 American Society of Hematology.

ASH Teaching Slides: Blood Cells

Normal blood smear. The four larger cells shown are called granulocytes, a type of white blood cell.

Photomicrograph of a Blood Smear

Figure 10.2

Cells you will see

• Red blood cells, erythrocytes, no nucleus, transport O2

• Platelets, deliver fibrin• White blood cells, leukocytes: complete

cells with a nucleus and organelles, defenders against disease,

Cell Differentiation and Lineages

• Granulocytes–Granules in their

cytoplasm can be stained– Include neutrophils,

eosinophils, and basophils

Figure 10.4

Eosinophiles

• Function: Phagocytosis of antigen-antibody complexes; allergens

• Release enzymes to weaken or destroy parasites such as worms.

• Appearance: Nuceus has 2 large lobes

Basophil

• Appearance:

• Function: Secretes Histamine,( a vasodilator)• Secretes heparin ( an anticoagulant)

Neutrophils

Appearance• Nucleus usually with 3-5 lobes in S-C shaped

array

Functions: Phagocytosis of bacteriaRelease of antimicrobial chemicals

Precaution: You need the right size drop to get a good smear

• Too large drop = too thick smear too many cells• Too small drop = too thin smear too few cells

• Thick and thin also depend on (de)hydration, hematocrit (anemia)

Peripheral Smear Preparation

2. The pusher slide is held securely with the dominant hand in a 30-45 deg angle.- quick, swift and smooth gliding motion to the other side of the slide creating a wedge smear

Peripheral Smear Preparation

Peripheral Smear Preparation

Precautions: Ensure that the whole drop of blood is picked

up and spread Too slow a slide push will accentuate poor

leukocyte distribution, larger cells are pushed at the end of the slide

Maintain an even gentle pressure on the slide – not doing so leads to ridges and valleys

Keep the same angle all the way to the end of the smear.

Peripheral Smear Preparation

Peripheral Smear Preparation

• Allow the blood smear to thoroughly dry – thicker will take longer

• Fix the blood smear with 70% methanol for 5 seconds; blot slide edge: let surface thoroughly dry

• Stain cells with Giemsa or Wright stain for 5’ - both are available; write in notebook which one you use – they color cells differently

• Dip slide in distilled water, 3 – 5 times• Blot edge; thoroughly dry

You are now ready to observe without and with microscope!!!

Feature of a Well Made Wedge Smear• Smear is 2/3 or ¾ the entire slide• Smear is finger shaped, very slightly rounded at

the feathery edge: widest area of examination• Lateral edges of the smear visible• Smear is smooth without irregularities, holes or

streaks• When held up in light: feathery edge should show

rainbow appearance• Entire whole drop of blood is picked up and

spread

• See shape, internal morphology, comparative size, count cells, identify disease conditions

Peripheral Smear Examination• Visual – no microscope: record in lab notebook– Accurately draw both smears– Measure length and width in mm– Assess overall quality, i.e., • Does it have a feathery edge? Fingerprint shape? • Quality of the staining –also assess at

• Microscopic:• 10x Objective record in lab notebook– Observe types and distribution of the cells – use systematic

approach, i.e., serpentine• where do you find red blood cells; are they easily distinguished,

i.e., separated• Check lateral edges for WBC (white blood cells) distribution; do

you see them distinctly• Quick sketch of 2 different fields; label and describe cells

Choose the best areas for assessment

• Systematic search• Back and forth serpentine or battlement

track patterns are preferred – move the slide mount

Peripheral Smear Examination• Microscopic:• 40x Objective this week and next

1. Determine correct area to view – cells spread apart but not too much

Optimal Assessment Area:1. RBCs are uniformly and singly distributed2. Few RBC are touching or overlapping3. Normal biconcave appearance

• Too thin

Peripheral Smear Examination

Too thick

Assessment at 40X

• Pick 5 fields that show different types of cells and sketch all cells; describe these cells in your own words

• Determine which cells you are seeing

Before you leave at 9:50

• Make sure your slides have your student ID#• Give to Younis; he will save for next week• Clean up your area

Final Assessment – Next Week

• Microscopic:• 100x objective, oil immersion