Materials and Methodology

A post mortem examination or autopsy is performed to determine the

cause of death. However, the cause of death might not always be
conclusive and include examination of any disease in prevalence aiding to
the death of the subject. Pathologists perform the postmortems under
regulations set by Human Tissue Authority and the Royal College
Pathologists. Autopsy results might take days to weeks for processing
from the time of subject’s death(Peres, 2017).

Our case study involved weighing the organs provided using large trays
with a ruler and balance to measure and weight the organs. The organs
are compared to reference ranges provided in the practical for any
abnormal observations in weight. Also, the organs are carefully identified
and for any physical damage like a scar or abrasion. After the organs like
heart, liver, right lung, left lung and right kidney are appropriately
identified, approximately 1 cm (centimeter) width and length with a 5
mm (milli meter) depth tissue is collected from each of them by a scalpel.
The tissue samples are fixed onto a glass slide for further testing using a
fixator like formalin. These six tissue samples are accompanied with a
request form and patient identifiers for histopathology testing. Swabs
from the six organs are also collected for microbiology analysis
accompanied with patient identifiers’ form.

Histology process included staining of heart, liver, heart, kidney and lung
with Haematoxylin and eosin (H & E) along with another section of liver
stained with Martius Scarlet Blue (MSB). The materials included and not
limited to are histoclear, Haematoxylin (Harris and Mayer’s) solution,
Eosin solution, Martius Scarlet blue stain, acidic alcohol: 1% HCl in
70 % ethyl alcohol, 1% acetic acid in water, DPX mounting medium,
coverslips, microscope, distilled water, Brilliant crystal violet solution,
phosphotungstic acid solution and Aniline blue dye solution. Table 1
gives an insight of these stains and their use in tissue diagnosis.
Table 1: H&E stain and MSB stain uses in pathology of tissues.
(Alturkistani, Tashkandi and Mohammedsaleh, 2015)

H & E staining MSB staining

Standard staining to identify
normal and pathological
architecture in tissues.
A blue/red intermediate shade is
produced on cellular and tissue
component in this staining.
Haematoxylin is a basic dye with
an affinity for cellular acidic
elements like nucleic acids.
Eosin is acidic dye exhibiting
affinity for basic elements like cell
cytoplasm.
Trichrome staining to identify
fibrin deposits, collagen
architecture and erythrocytes. Liver
pathology like cirrhosis, steatosis
and hepatitis can be observed.
Martius is a yellow small stain that
selectively stains erythrocytes when
combined with phosphotungstic
acid in alcoholic solution. Early
deposits of fibrin can also be
observed here. The
phosphotungstic acid blocks
subsequent staining of other tissue
structures.
Brilliant Crystal Scarlet is a red
medium sized dye and satins the
muscle and mature fibrin.
Aniline blue is a relatively large
molecule and stains collagen and
old fibrin.

The staining process is followed by a liver extraction practical for

toxicology testing was performed according to the guidelines provided.
Equipment and reagents used but not limited to tissue homogenate,
Gilson pipettes: 1 mL (milli litre) and 200 µL (micro litre), eppendorf
tubes, bench centrifuges, acetaminophen stock solution with 1 mg/mL
(milli gram/milli litre) concentration, ethyl acetate, phosphate buffer
solution (PBS) and hot plates with approximately 60 degrees Celsius
temperature are used.

A microbiology analysis for indication of presence of any pathogen as the

cause of death or presence as commensals is analysed by the studying the
morphology and ability of bacteria to grow on particular agar plates. The
materials and equipment used and are not limited to include crystal violet
stain, Gram’s iodine solution, safronin solution, light microscope, PBS
solution, centrifuge tubes, inoculation loops, laminar air flow chamber
and hydrogen peroxidase. The protocol is followed by the guidelines
provided and included isolation and identification of bacterial species
from blood cultures followed by gram staining: gram positive bacteria
retain violet colour due to presence of thick peptidoglycan layer and gram
negative bacteria stain pink (O'Toole, 2016), oxidase assay and catalyse
assay. The blood samples with the subject’s hospital number are cultured
on an agar plate after obtaining an appropriate gram stain results from
blood smears. Table 2 gives an insight of the agar plates to be used.

Table 2: Indication of agar plates in gram stain results

Gram Stain Agar Plates

Gram positive Cocci Blood, CLED (cysteine, lactose and
electrolyte deficient)
Gram positive Rods Anaerobe, CLED
Gram negative Cocci CLED
Gram negative Rods MacConkey, CLED

After obtaining appropriate results from gram staining, the bacteria are
cultured in appropriate agar plates and performed with catalase assay for
gram-positive cocci. Gram-negative rods, which were lactose non-
fermenters are subjected to oxidase assay. Positive oxidase test indicates
formation of blue colour and hence oxidase enzyme. Positive catalase
assay is indicated by effervescence. Both the tests were also performed in
inconclusive results of gram staining to obtain at least a potential
bacterial species.

Alturkistani, H., Tashkandi, F. and Mohammedsaleh, Z. (2015). Histological Stains: A

Literature Review and Case Study. Global Journal of Health Science, 8(3), p.72.

O'Toole, G. (2016). Classic Spotlight: How the Gram Stain Works. Journal of Bacteriology,
198(23), pp.3128-3128.

Peres, L. (2017). Post-mortem examination in the United Kingdom: present and

future. Autopsy and Case Reports, 7(2), pp.1-3.