U2 LAB: PBS PREPARATION Flashcards
Evaluation of peripheral blood film is an intergal part of?
CBC or hemogram
PBS is required for?
enumeration and checking of cellular elements
Microscopic examination aids in providing info regarding?
formed elements in blood
Reliability of information obtained depends on?
well-made and well-stained films
T/F: Smears must be prepared immediately as possible.
True
Blood sample
Anticoagulated EDTA blood
Stain used
Wright or Wright-Giemsa stain
Systematic approach:
- 10x
- 40x
- OIO
Materials needd:
- Glass slides, coverslips
- Pasteur pipette
- Anticoagulated blood
- Tube holder
- Pencil for labeling
- PPEs
- Canister with yellow bag
- Sharps container
- Oil cloth
Method 1
Two-Slide or Wedge method
Simplest and most common method for smear prep
Two-Slide or Wedge method
Procedures for Method 1
Size of drop ng blood
2-3mm in diameter
Procedures for Method 1
Drop of blood should be placed where?
1cm from the end
Procedures for Method 1
Fingers used to hold spreader slide
thumb and forefinger
Procedures for Method 1
Angle of spreader slide
30-45 degrees
Features of a Properly Made Blood Smear
The film must cover?
2/3 to 3/4 length
Features of a Properly Made Blood Smear
The film is what shape?
finger shaped (rounded at feather edge)
Features of a Properly Made Blood Smear
What edges should be visible?
lateral edges
Features of a Properly Made Blood Smear
The film is smooth without irregularities, like?
holes or streaks
Features of a Properly Made Blood Smear
When slide is held up to light, feathery edge should have a?
rainbow appearance
Portion observed in feathery edge
Zone of morphology
Unacceptable Peripheral Blood Smears and its Causes
- chipped or rough edge
- hesitation in forward motion
- pushed too quickly
- drop of blood was too small
- not allowed to spread across the width of slide
- dirt or grease
- uneven pressure
- time delay
T/F: Hematocrit levels can be attributed to smears.
True
Low hematocrit levels
Too thin
High hematocrit levels
Too thick
Reminders once you have created the smear
A good film includes a?
thick and thin portion, with gradual transition
Moving the pusher slide forward too slowly accentuates?
poor leukocyte distribution (pushing larger cells at very end of film)
The faster the film is air dried,
the better is the spreading
Slow drying causes?
formation of artifactual cell
Slide may be labeled on the frosted end or directly on the thicker end using?
lead pencil
The thickness of the film can be adjusted by?
changing the angle or speed of spreading
This can cause erroneous result in preparing the smear
high lipid content
Older technique that is used for making bone marrow aspirate smears.
Two-Coverslip or Two-Coverglass method
Method 2
Two-Coverslip or Two-Coverglass method
Recommended coverslips for Method 2
No. 1 or 1 1/2 cover glasses 22mm
Quality coverslip smear will appear similar to a?
thumb print
Making smears on coverslips require?
manual dexterity
Advantage of Method 2
- excellent leukocyte distribution
- minimal amount of bone marrow is needed but many smears are made
Procedures for Method 2
Hold one coverglass on adjacent corners with?
Thumb and index finger
Procedures for Method 2
Blood must form a?
16-sided figure (letter B)
Procedures for Method 2
Before blood spreads, seperate coverglass by?
rapid, even, horizontal and lateral pull
Method 3
Spinner Method
This method would create a uniform blood film.
Spinner Method
Easy handling of wedge slide and uniform distribution of cells can be made with special types of centrifuges known as?
spinners
Spinner slide produces film in which?
all cells are separated (monolayer) and randomly distributed
Smears prepared using Method 3 have what that can be easily identified at any spot in the film?
white blood cells
On wedge smear, disproportions of ____ occur at tip of feather edge
monocytes
____ are seen just in from the feather edge
Neutrophils
___________ may occur at lateral edges of the films
Neutrophils and Monocytes
This is an automated slide making and staining system.
Sysmex SP-1000i
Automated Smear Preparation is dependent on?
hematocrit values
Film for automated smear prep is produced every ____ and with what?
30 seconds, printed label
Printed label for Automated Smear Prep contains?
- Patient name
- Number
- Date of preparation
This prepares slides automatically based on orders received from the LIS
DxH Slidemaker Stainer II Cellular Analysis System by Beckman Coulter
DxH Slidemaker Stainer II Cellular Analysis System by Beckman Coulter
This creates and stains smears from?
- capped whole blood tubes
- open-top tubes
- pediatric whole blood tubes
DxH Slidemaker Stainer II Cellular Analysis System by Beckman Coulter
How many slides can it create from a single 90uL sample?
4 slides
DxH Slidemaker Stainer II Cellular Analysis System by Beckman Coulter
How many slides can it create from a single sample presentation?
12 slides
DxH Slidemaker Stainer II Cellular Analysis System by Beckman Coulter
This creates multiple stain smear slides according to?
any parameter results
Wright stain is composed of?
- Eosin
- Complex mixture of (Thiazines) including methylene blue, azure blue
This staining technique is certified by the Biological Stain Commission and is commercially available as solution or powder.
Staining Jar or Dip Method
T/F: Polychrome stains contain Eosin and Methylene blue.
True
Heparin causes?
blue background
Oxalate and Fluoride causes?
distortion of cell morphology
Reagents in staining dip or jar method is found in?
coplin jars
Solution 1
Methanol, 30 seconds
This will serve as fixative.
Methanol
Solution 2
Eosin, 6 seconds
Free eosin is acidic and stains basic components such as?
Hemoglobin or eosinophilic granules (red)
Solution 3
Methylene blue, 4 seconds
Free methylene blue is basic and stains acidic cellular components such as?
RNA (blue)
Solution 4
Buffer solution/aged distilled water, 45 seconds
Buffer solution
0.05M sodium phosphate (pH 6.4)
Aged distilled water
Placed in glass bottle for at least 24 hours; pH 6.4 to 6.8)
Slide must dry in a?
tilted position
Color of RBCs
pink to salmon
Color of Nuclei
dark blue to purple
Cytoplasmic granules of Neutrophils
lavender to lilac
Cytoplasmic granules of Basophils
dark blue to black
Cytoplasmic granules of Eosinophils
red to orange
Area between the cells should be?
colorless, clean, free of precipitated stain
Slides must be prepared within how many hours following blood collection?
2 to 3 hours
Oxidation on staining causes development of?
Formic acid
Other romanowsky-type stains:
- Giemsa
- Leishman’s
- Jenner’s
- May-grunwald
- MacNeal’s