Unit #8: Cytology- Abnormal Fluids

What is transudate

Fluid substance that has passed through a membrane or has been forced out of a tissue

Why does transudate occur and what does it look like

• due to increased pressure in tissues or low amounts of protein (low oncotic pressure) to keep fluid in vessels

• Clear watery fluid

Define exudate

Fluid w high content of protein and cellular debris that has escaped from blood vessels & has been deposited in tissue or on tissue surfaces

Why does exudate happpen

Inflammation (pus)

Does transudate or exudate often clot?

Exudate clots

Transudate does not clot

What is exudate composed of

Protein and cells

What is transudate composed of

Filtered plasma w little protein and no cells

What is a chylous effusion

• milky fluid produced during digestion that enters lymphatics

• Uncommon

3 key tests to differentiate body fluids

• protein content

• Nucleated cell count

• Modified transudate have higher numbers of cells and protein due to being more chronic

When measuring protein content what do we use?

Refractometer

Example of a table comparing transudate and exudates

What are some conditions that would produce transudates (4)

Possibly:

• hypoproteinemia, live disease, heart disease, tumor

Possible conditions that would produce modified transudates (3)

Possibly

• CHF

• Abdominal tumour

• Liver disease

What are some conditions that may produce exudates 5

Possibly

• bleeding conditions

• Bacterial infection in cavity

• Chyloabdomen/chylothorax

• Urine and bile peritonitis

• Pancreatitis

If a cytology shows that the fluid is due to inflammation, it can lead to 1 of 2 thigns

Septic or non septic

If the cytology is non inflammatory, it can lead to 2 things

Non inflammatory= neoplastic (tumour) or non-neoplastic

If neoplastic (tumour)= benign or malignant

How much fixative should tissues be stored in when handling specimen

1 part tissue to 10 parts fixative

*10% formalin or alcohol

5 steps for specimen handling

1. Check before sample is collected for necessary info

2. Handle specimen appropriately (gloves)

3. Use buffered fixative for tissues

4. Don’t allow tissues to dry out

5. Seal containers

How do we dispose of formalin (formaldehyde)

Formalex

• liquid that can be mixed w formaldehyde so it can be disposed into sewer system

What must be labeled on a sample? (5)

• owners name

• Patient name

• Sample id (kidney biopsy)

• Date collected

• Method of collection

4 important instructions for tissue submissions to the lab

• samples cant be more than 5-10mm thick

• Multiple skin biopsies should be submitted in seperate containers

• Protect cytology slides from formalin fumes

What should tissue submissions of tumors or skin lesions include

• abnormal tissue

• Border between normal and abnormal tissue

• Adjacent normal tissue

*do NOT rinse tissue w water

What do we stain tissue impression smears with if vet wants margins checked

Indian ink

What is a synovial fluid cytology submission called

Joint capsule aspiration

Lymphocytes characteristic

• smaller than neutrophil

• Large nucleus

• Most common wbc in cattle

• 2nd most commmon in all other species

8 most common cell types that may be seen in fluid

• rbc

• Neutrophil

• Lymphocytes

• Monocyte/macrophage

• Plasma cell (B cells)

• Eosinophils

• Mast cells

• Mesothelial cells

2 reasons for large numbers of RBC in fluid samples

1. Recent hemorrhage

• platelets present

2. Associated w disease process

• Erythrophagocytosis may be present

4 names of changes to the nucleus in neutrophils in fluid samples

• Pyknosis

• Karyorrhexis

• Karyolysis

• Hypersegmentation

What is pyknosis

Indicates non toxic enviroment

Slow progressive cellular death

What is karyorrhexis

Rupture of nucleus in neutrophils

• weakly toxic environment

What is karyolysis

Nucleus on neutrophil stains pale

• indicates rapid cell death

• Highly toxic environment

What is hypersegmentation in neutrophils

More than 5 lobes in the nucleus

• chronic pus forming lesions

5 toxic changes occurring in neutrophils

• cytoplasmic basophil is

• Toxic granulation (blue stain granules)

• Toxic vacuolation (appear to have foamy cytoplasm)

• Dohle bodies In cytoplasm (light blue rod shape)

• Nuclear immaturity

If you see toxic changes in neutrophils, what should we also loook at?

Bacteria in cytoplasm- if present it’ll indicate septic inflammation vs non-septic inflammation

Is it common to see neutrophils in transudate

No- sometimes low #s

Possible pyknosis or hyper segmentation of nucleus

Are neutrophils common in exudate

• usually large #s present

• Usually indicates acute inflammatory condition

When do lymphocytes most commonly appear

Chronic inflammatory exudates

How long after acute inflammation starts do monocytes/macrophages appear

8-10hrs

What do plasma cells (b cells) indicate

Chronicity of fluid sample

What do increased numbers of eosinophils indicate

• allergic reaction

• Parasitic reaction

What do mast cells indicate

• release histamine

• During inflammatory and allergic reactions

What is the function of mesothelial cells

• line bodies serous cavities (pleural, peritoneal, pericardial)

• Easily mistaken for tumour cells

6 tests that should be performed on synovial fluid sample

A. Color and clarity

B. Viscosity

C. SG

D. Total Protein

E. WBC count

F. Cytology

What does viscosity testing on synovial fluid indicate

• evaluates hyaluronic acid

• Place drop of synovial fluid between

What is cytology?

Microscopic examination of cells from tissues, organs and body fluids

• FNA

• Swabs

Name an advantage of performing cytology (several)?

Rapid turnaround time

Name a potential risk/consideration of choosing to perform cytology?

Rarely, seeding of infection or neoplastic cells from the primary lesion to the surrounding healthy tissues during collection may occur.

What are 3 possible reasons for a poor-quality/non-diagnostic cytology?

1. hemodilution of the sample

2. the actual aspiration technique

3. slow drying of the slide

What does histopathology refer too?

microscopic examination of a piece of tissue to evaluate the cells and architecture together to help identify or rule out disease.

• biopsy

• more invasive

• takes longer

True or False: Cytology provides a more definitive general disease diagnosis than histopathology

False, cytology does not relay information about the tissue architecture, which may affect how the cells are interpreted.

Ture or False Cytology is better for the diagnosis where identification of an individual cellular morphology is important.

True

Name three types of inflammatory cells you may see on a cytology smear (of many).

1. Neutrophils

2. Macrophages

3. Multinucleated giant cells

True or False: a “Purulent” inflammation contains predominantly macrophages.

False- contains predominantly neutrophils

True or False: Suppurative (purulent) inflammation may occur secondary to trauma or chemical injury (where no infection is present).

True

When evaluating a cytology sample for bacteria, to know if it’s causing the infection or not, where is an extremely important place to look?

Intracellular bacteria (look inside neutrophils) to confirm that the bacteria are pathogens.

What cells are predominantly found in each type of inflammation?

1. Granulomatous or Pyro-Granulomatous inflammation: Mixture of macrophages and neutrophils

2. Purulent inflammation: Neutrophils

What are two methods for acquiring a cytology sample (of many).

1. Fine needle aspiration (FNA)

2. Impression smear

What are two things you should do after collection to prepare a fluid sample to send out for cytologic evaluation?

1. Make direct smears using a sterile swab onto a glass slide immediately after collection.

2. Place small amount of sample in both a red top and a lavender top tube for culture and cytological evaluation.

What are three things you should do/ensure when you are preparing a solid tissue sample cytology for cytology

1. Blot excess fluid before doing smears

2. If submitting the sample for histopathology, make sure that you do not stain the sample

3. 3. If submitting slides with tissues which have been fixed, ensure there is no leakage of formalin fumes. *formalin/glutaraldehyde fumes may alter the staining characteristics of the smear.

What are the two common stains used for cytology?

1. Romanowsky stain

2. New methylene blue

Transudate

Appearance: Clear, thin, pale yellow to colourless

Protein content: Low, <2.5 g/dl

Nucleated cell count: <1500/uL

Cell type: Lymphocytes, occasional neutrophil, some mesothelial cells, RBCs

Coagulation of fluid: None

Microorganisms: Absent

Modified transudate:

Appearance: Slightly turbid, yellow to red/reddish brown/ milky,

Protein content: 2.5 - 7.5 g/dl

Nucleated cell count: 1500/uL - 7000/uL

Cell type: Mesothelial cells, RBCs, neutrophils, macrophages

Coagulation of fluid: None to slight

Microorganisms: Absent

Exudate

Appearance: Cloudy/opaque, variable colour depending on content: red, creamy, amber or other

Protein content: >3.0 g/dl

Nucleated cell count: >7000/uL

Cell type: Neutrophils, mesothelial cells, usually RBCs, lymphocytes, macrophages.

Coagulation of fluid: Usually clots

Microorganisms: Often demonstratable

Pyknosis:

Small, round, dense staining nucleus

Karyorrhexis:

Rupture of nucleus into fromless granules

Karyolysis:

wollen, ragged nucleus that stains palely

Hypersegmentation:

>5 lobes on the nucleus

What are the 5 cytoplasmic changes that indicate immaturity/toxic change in cells?

1. Cytoplasmic Basophilia

2. Toxic Granulation

3. Toxic vacuolation

4. Dohle Bodies within the cytoplasm

5. Nuclear immaturity

True or False: There will be low numbers of neutrophils in transudate fluid.

True

True or False: With acute and chronic inflammatory exudates lymphocytes will not be seen.

False

What is the general ratio rule when it comes to adequate fixation of tissue with formalin.

10% buffered formalin is 1 part tissue to 10 parts buffered formalin.

True or False: Formalin is not carcinogenic.

False

What are five things you should put on your sample label?

1. Owner name

2. Patient name

3. Sample identification

4. Date

5. Method of collection

True or False: A sent to the lab should be no more than 5-10mm in size

False- 5-10 mm in thickness, length and breadth, can be several centimeters.

True or False: If a tissue sample is covered in blood or exudate, it is fine to rinse it off with water before examination

False- don’t rinse with water, use fixative or saline