Sampling Techniques Flashcards
What is cytology?
It is the study of cell number and type in a tissue mass of fluid accumulation to investigate its cause (usually inflammation or neoplastic).
Name three things cytological examination should enable?
- Differentiation of different fluid types e.g. transudate vs exudate
- Differentiation of type of inflammation e.g. eosinophilic, neutrophilic, granulomatous (fungal)
- Detect the presence of neoplasia.
What are the 4 scenarios which might lead to a false negative?
- The tumour might not exfoliate very well
- Might not have got cells from the right sample site
- Extensive inflammation around the tumour can look like ordinary inflammation
- Tumour might not be well enough differentiated to allow an accurate diagnosis.
What might cause a false positive?
Dysplasis can mimic neoplasia and occurs in inflammatory disease.
What is histopathology?
The study of changes in tissues caused by disease.
What is FNCS and how is it performed?
Fine needle capillary sampling - no suction- needle with no syringe attached
What is FNA and how is it performed?
Fine Needle Aspirate - minimal suction
When would you use FNA?
Cysts or failed FNCS - needle and syringe attached
What can we determine from fluids?
Their classification and tissue of origin.
What is a lavage and where could it be used?
Saline is used to wash nasal, bronchoalveolar, urinary bladder mucosa and the resulting fluid is collecting by collecting of the immediate lavage.
What tubes would you use to collect lavage samples?
Split between EDTA and sterile tubes.
When do lavage samples need to be processed?
As soon as possible.
What is the most sterile collection method of urine?
Cystocentesis
Is a little body cavity fluid normal?
A little is normally present, however too little to collect, unless in horses.
What causes profound amounts of cavity fluid?
Hypoproteinaemia.
What is BAL?
Bronchoalveolar lavage
How is BAL performed?
Place catheter into the trachea, add saline and quickly aspirate. The fluid is then centrifuged and a smear is made from the centrifuged sediment.
What is bronchoscopy?
Viewing with a camera - cells can be removed with a tiny brush
What is a problem with broncho-alveolar lavage?
Might pick up bacteria from the pharynx.
What are the 4 basic tests applied to fluid samples?
- Appearance of fluid
- Cell type
- Total protein content
- Total nucleateWd Cell Ceount (TNCC)
What are the two methods to prepare a slide for cytological examination?
- ‘wedge’ method
2. Flat-Slide method
What have you got to be careful of when preparing a smear?
not to CRUSH the cells
If the fluid is turbid, what should you do?
make the smear directly on the glass slide.
If the sample fluid is clear, what should you do?
Centrifudge the fluid and smear the sediment (might be a low cell count).
What type of centrifuges are used to yield cells when the cell count is low?
Cytocentrifuges.
Once you have got your smear, what should you do?
Air-dry rapidly and stain
What happens if you don’t dry rapidly?
All sorts of artefacts
For how long and what speed should a bench centrifuge be used?
5 mins at 500-1000rpm
What can biopsy and necropsy tissues be used for prior to sticking in formalin?
To make impression smears for a rapid preliminary diagnosis.
What is the benefit of taking an impression smear before sending the sample of for histopathology?
Histopathology can take weeks and impression smears give us a preliminary idea.
What is a rule regarding cytology and formalin
KEEP CYTOLOGICAL SAMPLES AWAY FROM FORMALIN FUMES.
What effect does formalin have on a cytology smear?
Ruins it.
What should you NEVER do with wet slides following FNA and why?
Never put wet slides in the carrier box as they will start to rot.
What 5 things should be included with the smears?
- Description of the mass
- Location
- Size
- Growth rate
- Texture
Name 5 types of Romanowsky blood stains?
- Diff-Quick
- Dip-Quick
- Wright’s
- Giemsa
- Leishman
What stain would be used for Mast cells?
Toluidine Blue
What stain would be used for mucin?
PAS - periodic acid schiff
Name 4 stains that could be used if microorganisms were present?
- Gram
- Ziel-Nielsen
- Fontana
- PAS
What are the 6 types of effusion categories?
- Exudate
- Transudate
- Haemorrhage
- Lymphorrhage
- Organ rupture
- Mixture of above
What is g/dl x 10?
g/L
What is transudate?
Excessive diffusion of plasma water from vasculature (transudation).
Name two reasons for increased transudate?
- Increased hydrostatic pressure e.g. hypertension (Na+ retention, increased alveolar capillary pressure)
- Decreased COP (hypoproteainaemia, hypoalbuminaemia)
What is the second reason other than increased transudation, which causes transudate?
- Accumulation
What causes accumulation?
- Impaired lymphatic drainage e.g. congestion in vena cava leading to venous congestion
What are the two causes of transudate?
- Increased transudation
2. Accumulation e.g. poor lymphatic drainage
Should vasculature be low or high in protein and when does this change?
Normal, healthy vasculature should be impermeable to protein therefore transudate should be low in protein. Inflammation makes vessels more leaky therefore transudate protein content will increase.
What are the two causes of ‘protein poor’ transudates?
- decreased COP (hypoproteinaemia).
2. Portal hypertension e.g. increased Hydrostatic pressure e.g. crirrhois.
What are the causes of protein ‘rich’ transudates?
Come from interstitium, not from vessels.
This varies by organ - 2g/dl in subcutis, 6g/dl in liver.
- portal hypertension e.g. CHF (post-sinusoidal).
What is another term which describes ‘protein rich’ transudate?
Modified transudate.
What is modified transudate?
A transudate which has been modified by the presence of protein or cells.
Why is modified transudate not commonly used as a descripition term?
Because it does not assist with differentiating the origin of the transudate.
What does normal transudate contain?
Not many cells or protein
What does exudate contain?
Lots of cells and protein?
Modified transudate?
This is a grey area between transudate and exudate. Somewhere in between.
What is an exudate?
Exuding or leaking out of pores.
Why does exudate occur during inflammation?
Increased vascular permeability.
What does exudate contain?
Lots of cells and proteins
What kind of cells does exudate contain?
Inflammatory cells e.g. macrophages, neutrophils, lymphocytes and eosinophils.
Why does FIP contain lower cell counts?
Because this is due to vasculitis rather than pleuritis.
What is characteristic of mesothelial cells?
They have a very basophilic cytoplasm with variable cytoplasmic ratio.
Is it abnormal for mesothelial cells to be multi-nucleated?
No, they might have clumped chromatin and prominent nucleoli.
What else helps to determine cells as mesothelial cells?
A pink ring surrounding the cells.
What is a cytological low protein reading?
<20g/L
What is a low nucleated cell count reading?
<5x10(9)/L
What might mesothelial cells present when there is irritation or inflammation present?
Become dysplastic.
Name 3 examples when you would find exudates?
- inflammation
- Necrosis (could be within tumours)
- Infection
What protein reading would you commonly find in exudate?
> 35g/L
What nucleated cell content would you commonly see with exudate?
5-25 x 10(9)
What cells would you commonly see with exudate?
RBCs, mixed nucleated cells, maybe dysplastic cells?
Why might tumours produce inflammation?
Because they have a ‘foreign body effect’. There might also be some areas of necrosis.
what is the cytology of haemothorax?
Appearance: Blood (but does not clot)
PCV: variable >0.03g/L
Protein: >20g/L
Nucleated cells: variable but similar to blood 5-15x 10(9)/L
Cell types: RBCs, plateles, mixed nucleated and mesothelial cells, possibly hemisiderophages (alveolar macrophages)
What is the cytology of uroperitoneum?
Appearance: Serosanguineus, might smell of urine, especially if heated.
Protein:10-30g/L
Nucleated cells: Low at first, progresses to 5-15 x 10(9)/L
Creatinine and potassium: >plasma if recent or ongoing
Cell types: RBCs, Mixed nucleated cells e.g. neutrophils, macrophages, mesothelial cells
What is the diagnostic cytolosy of chyle/ chylous effussion?
Appearance: milky, white or pink
Protein: >20g/L
Nucleated cells: 5-20g/L
Cell types: Varies with the age of lesion but mainly mature lymphocytes at first.
Name some reasons for the cause of chylothorax/chylous effusion?
Trauma, spontaneous rupture of the thoracic duct.
Heart failure, cardiomyopathy, pericardial effusion, neoplasia, lymphangectasia, obstruction of minor lymphatics by chronic inflammation.
What is common in cats?
Non-thoracic duct origin chyle in the thorax due to cardiomyopathy, diaphragmatic hernias etc.
What is the turbidity of chyle?
++++
What is the turbidity of non-chyle?
+++
What is the triglyceride content of chyle?
> plasma
What is the triglyceride content of non-chyle?
What is the cholesterol content of chyle?
what is the cholesterol content of non-chyle?
> plasma
What is the chol:tri ratio of chyle?
low
What is the chol:tri ratio of non-chyle?
high
What are the predominant cells in chyle?
Lymphoctes
What are the common cells in non-chyle?
Neutrophils and macrophages
What occurs after centrifugation of chyle?
Turbidity remains
and centrifugation of non-chyle?
Turbidity remains
What happens when ether and NaOH is added to chyle?
Clears
What happens when ether is added to non chyle?
turbidity partly clears.
What are the 5 things that are examined wit synovial fluid?
- gross appearance
- viscosity (decreases with disease)
- total protein count
- total nucleated cell count
- cytology
What is the mucin clot test, what is added and what is normal?
This is the addition of acetic acid. there is formation of a big clot with healthy synovial fluid and not much clot formation with diseased synovial fluid.
What does a tight clot on the mucin test show?
Good viscosity
What should total protein of synovial fluid be in dogs and cats? g/L
<25g/L
What should total protein of synovial fluid be in cattle and horses? g/L
<20g/L
What should total nucleated cells be in cats and dogs in synovial fluid (x10(9)
<3
What should total number of nucleated cells be in synovial fluid of horses? (x10(9)?
<0.5
What should total number of nucleated cells be in synovial fluid of cattle? (x10(9)
<1
What structure do cells take on synovial fluid smear?
Cells form proteinacious rows
What indicates inflammation of synovial fluid?
If >10% mononucleated cells.
What is the total protein, TNCC and cytology of degenerative joint disease?
Total protein normal, TNCC normal, cells normal, although occasionally might see dysplastic synoviocytes.
What is the total protein, TNCC and cytology of inflammation-non septic?
Common in cats and dogs (immune-mediated)
Total protein: increased
TNCC: increased
Cells: neutrophils will predominate
What is the total protein, TNCC and cytology of inflammation-septic?
Common in horses and ruminants.
protein increased, TNCC increased, Cells, mainly neutrophils.
What cells are normally predominant on a BAL?
Ciliated columnar epithelial cells.
<10% leukocytes
up to 20% eosinophils in cats
What suggests the presence of oropharyngeal contamination?
Squamous epithelial cells and simonsiella bacteria.
What do you look for on bone marrow examination?
- Cellularity (increased, decreased, normal)
- Cell lineages (myeloid (WBC), epitheloid (RBC), megakaryocytes (platelets))
- Neoplastic cells
- Parasites
