BIOT 309: BLOOD SMEAR PRACTICAL

Feb., 2013

Microscopic Views

Bird Blood Horse Blood Cat Blood Dog Blood Human Blood Fish Blood Frog Blood Snake Blood

ASH Teaching Slides: Blood Cells

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

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

Photomicrograph of a Blood Smear

Figure 10.2

Cells you will see

• • •

Red blood cells, erythrocytes, no nucleus, transport O 2 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:

Histamine,( a vasodilator) Secretes Secretes heparin ( an anticoagulant)

Neutrophils

•

Appearance

Nucleus usually with 3 5 lobes in S-C shaped array

Functions:

Phagocytosis of bacteria Release of antimicrobial chemicals

Peripheral Smear Preparation

• • Precaution: You need the right size drop to get a good smear Too large drop = too thick smear cells  too many 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

• • • • • 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 distributed 2. Few RBC are touching or overlapping 3. Normal biconcave appearance

Peripheral Smear Examination

• Too thin 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