Histology and cytopathology

Question 1

What are the tools available for histopathologists

Answer 1

Histopathologists (interested in tissues):
Biopsies
Resection specimens
Frozen sections
Post-mortems

Question 2

What are the tools available for cytopathologists

Answer 2

Cytopatholgists (interested in cells):
Smears
Fine needle aspirates

Question 3

Summarise the purposes of histopathology

Answer 3

Asess spread and type of cancer

Monitor treatment in chrons – grade inflammation

Biopsies- percutaneously, endocscopy
Resection- from colon, liver or pancreas- any organ except brian

Question 4

Describe biopsies

Answer 4

Is it normal?
Is it inflamed and, if it is, what is the cause?
Is it cancer and, if it is, what type is it?

Impoertant to say type of tumour- for treatment

Question 5

Describe resection specimens

Answer 5

How far has the cancer spread?
Is it all out?
A subtotal colectomy is where the surgeon removes most of the colon but is able to leave part of the left side of the colon called the sigmoid colon. The surgeon joins the small bowel to the remaining bit of large bowel (sigmoid colon- so usually have diagnosis before hand
take marigns of resection- to see where tumour has spread- IBD- look for inflamed margins- as in reconstruction- make anastomiss between two parts of bowel- if infklammed – won’t work

Question 6

Describe frozen section

Answer 6

Rapid diagnosis
Is it cancer?
Is it all out?
Is there something else going on?

Still in theatre- want to know if its cancer- want to remove all of it- to know if margins are clear- shows they took everything out
May find lymphoma
Need to do as quickly as possible

Question 7

Describe post-mortems

Answer 7

Hospital post-mortems

Coroner’s post-mortems

Hospital- requested by clinician
Coroner- law- suspicious death

Question 8

Summarise how samples are obtained

Answer 8

Specimen must be properly labelled
Fix in formalin
Embed in paraffin wax
Cut sections

Question 9

Describe how samples are obtained

Answer 9

Label pot with patient details and where biopsy is from

Tissue in formaline- helps tissue to fixate- easier to examine tissue- 10% formaline- slow penetration in tissue- if tissue not fixed- leave in pot for longer

Microtome- to cut section from paraffin wax- to get slide with section- stain H and E (routine stain)- pink, pureple colour

Question 10

What can we do with the sections

Answer 10

Stain e.g. gram, ZN.
Identify specific antigens using antibodies = immunohistochemistry
Carry out molecular tests

Zn from TB- background blue TB red

Question 11

Describe cytopathology

Answer 11

Looking at individual cells not tissues
Used for fine needle aspirations

Obtained by fine needle aspiration or smears (biopsies)

Question 12

How long do histopathology results take to reach the clinician

Answer 12

Frozen section: 30 minutes
For biopsies: 2-3 days
For resection specimens: 5-7 days

Question 13

Describe Kaposi’s sarcoma

Answer 13

Billy’s skin biopsy shows Kaposi’s sarcoma
This is an HIV AIDS defining disease

Vasculat tumour- can occur in skin, also involve internal organs- especially bowel

Immunocytochemistry for CD31 (brown) to show vascular tumour infiltrating collagen bundles

A fine needle aspiration of one of the enlarged nodes revealed a mixed cell population

The diagnosis is of a reactive lymphadenopathy

Question 14

What is meant by pathology

Answer 14

Pathology is the medical science and specialty practice that deals with all aspects of disease, but with special reference to the essential nature, the causes, and development of abnormal conditions, as well as the structural and functional changes that result from disease processes – more simply it is the study of disease. The literal translation of pathology from Greek (pathos, -logos) is “the words of suffering.” Pathologists study: the causes of disease; how various internal and external derangements or injuries affect certain cells and tissues; the progression of disease in the human body (its pathogenesis); how disease is manifest (its clinical expression and the lesions produced); and methods for monitoring disease progression.

Pathology provides a scientific foundation for clinical medicine and serves as a bridge between the basic sciences and patient care

Question 15

Describe the purpose of histopathology

Answer 15

Histopathology encompasses surgical and autopsy pathology and histopathologists make diagnoses on tissue – biopsy material, surgical specimens removed at operation, make rapid diagnoses when necessary at operations by use of “frozen sections”. The autopsy (post mortem) is performed to determine the cause of death of a patient, explain why treatment was unsuccessful, to show the spread of disease and to educate doctors regarding disease processes.

Question 16

Describe the role of cytopatholgists

Answer 16

Cytopathologists make diagnoses on cells – cellular specimens – sputum, body fluids which contain cells, cervical smears and tissue obtained by fine needle aspirate (FNA). This is a less invasive technique than obtaining tissue for a biopsy and thus has advantages to the patient.

Question 17

Why are biopsies taken

Answer 17

Biopsies are taken for numerous reasons. Common conditions that require biopsies are skin lesions (inflammatory and suspected tumours). If a tumour is suspected clinically this may be removed intact, with a margin of normal skin to ensure excision of the tumour. However a rash may simply require a punch biopsy to determine the nature of the inflammation.

Question 18

Describe biopsy of different tissues in the body

Answer 18

Endoscopy enables the clinician to view the gastrointestinal tract. Biopsies are commonly taken from the stomach to exclude for example cancer, the duodenum to exclude coeliac disease and the large bowel to confirm cancer or diagnose inflammatory bowel disease.

Bronchoscopy allows a view of the trachea and bronchi and biopsies of suspected tumours or inflammatory lung conditions can be taken.

Liver biopsies can be performed under imaging guidance for tumours or for diagnosis of liver disease.

Renal biopsies are taken to determine the nature of glomerulonephritis and other renal disease.

Question 19

Describe the benefits of cytopatholgy

Answer 19

Cytopathology is very helpful in diagnosis of lung tumours and is non invasive.

Cytopathology can also aid diagnosis in endoscopy when brushings of the tumour can be taken. It is used in cervical and breast screening. Fluids such as urine and ascities can be examined for cells.

Surgical specimens removed at operation are all sent to Histopathology for diagnosis and to determine if a tumour is completely excised.

The histopathologists and cytopathogists will generate a report of the examination of the specimen, which is sent to the clinician responsible for the patient. It is important that the report gives a conclusive diagnosis and a statement of tumour excision and prognostic features – for example if surrounding lymph nodes are involved or the tumour spread on to the peritoneum.

Question 20

Describe the purpose of formalin

Answer 20

Specimens for histopathology (biopsies and whole tissue) must be “fixed” in formalin, a preservative that stabilises protein bonds and prevents autolysis. The tissue then must be processed, sections cut and stained and the histopathologist must write the report.

Question 21

Describe the time scales of the different specimens

Answer 21

For large specimens, allowing for overnight fixation or longer if the specimen is large or fatty, this process should take 2-3 days. Small biopsies can be processed in a day and if a rapid
diagnosis is required then rapid processing takes 4-5 hours.

A very rapid diagnosis may be required during an operation – is this a tumour? Is the margin of excision adequate? Is this lymph node involved? Have I got the parathyroid?

Frozen section tissue is received in the laboratory without any fixative. A small sample is selected and frozen rapidly. A thin section is cut on a microtome contained in a refrigeration cabinet, fixed rapidly and then stained with haematoxylin and eosin. An answer can be given by phone within 20 minutes.

Question 22

Describe the process of acquiring specimens

Answer 22

All specimens are allocated an identity number and logged in to the computer.

During the daily cut up of specimens the pathologist or biomedical scientist selects and describes the tissue samples to be examined. Some specimens (for example biopsies) are processed whole, while larger specimens (e.g. mastectomies, colonic specimens) have a few selected pieces removed. Each selected piece of tissue is placed in a small perforated plastic container with a lid.

This plastic cassette receives the laboratory number. These tissue blocks are processed to paraffin wax. This small sample of the tissue must have the water extracted by dehydration through alcohols to be impregnated with paraffin wax that allows the block to be cut to thin sections using a microtome and the section mounted on glass slides. Sections are usually cut at around 5 microns thick – a micron is one thousandth of a millimetre!

The number of blocks cut every day varies from 50-400 and several sections may be taken from certain blocks – levels through the tissue to extract maximum information. The sections are rehydrated and stained with haematoxylin and eosin (H&E). Haematoxylin stains nuclei blue and eosin the cytoplasm pink. The sections are then viewed using the microscope.

Question 23

Describe some additional stains available

Answer 23

Most diagnoses can be made by simple H&E staining, however additional stains can help diagnosis. According to the dyes or chemicals used it is possible to stain certain tissue components selectively. For example, silver nitrate solutions may be used to identify melanin pigment, fungi, calcium deposits and certain types of fibres. Other dyes are used to show glycogen, mucins and types of structure within the tissues. It is also possible to demonstrate tuberculosis bacilli by the Ziehl-Neelsen method and bacterial infections by Gram’s method. However for difficult tumors immunocytochemistry techniques are needed which use antibody systems to label antigens within tissues. Thus epithelial, mesochyme, nerve, and prostate tissues can be specifically identified.

Question 24

Why are autopsies performed

Answer 24

Autopsies may be performed by authority of the Coroner (suspicious/ unknown cause of death) or by permission of the relatives of the decreased – the questions that can be answered include:
What happened as a terminal event?
Was the treatment successful but complications occurred?
Where did the cancer originate?
Was my diagnosis correct?

Question 25

Describe the use of cervical smears

Answer 25

Cervical smears are prepared preserved (fixed) with alcohol, stained by a special stain called Papanicolaou and permanently preserved. Most of the preparation methods within the laboratory are automated. Preparatory work is carried out by well-trained and experienced Medical Laboratory Scientific Officers. Slides are read by trained screening technical staff and abnormalities found checked by consultants.

Cervical smears are performed to pick up cervical cancer as part of the screening programme – Screening only has value if these (and other) criteria can be met:

Test is easy and non invasive
High take up in the population
A significant number of cases can be detected
Something can be done about the disease

Question 26

Describe the problems with cervical smears

Answer 26

However there are problems with for example, cervical screening:
Failure to obtain a satisfactory sample (smear)
Failure to make a proper smear
Poor staining
Poor interpretation