Methods of Diagnosis of Corona Virus

1. Diagnostic tests
for
Coronavirus disease 2019
(COVID-19)
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2. What is origin and How it spread?

3. What is COVID19?
COVID-19 – Coronavirus disease-19
A respiratory illness caused by severe acute respiratory syndrome
coronavirus 2 (SARS-CoV-2)
Corona has crown-like spikes on the outer surface of the virus
Coronaviruses are minute in size (65–125 nm in diameter)
Single-stranded RNA as a nucleic material, size
Why named Corona Virus?

4. What is structural characteristics?
Spike protein: S - proteins are functionally
divided into two subunits (S1 and S2).
S1 domain: Responsible for receptor
binding to receptor on target cell
S2 domain: Responsible for fusion
N protein : plays an important role in viral
pathogenesis, replication, and RNA
packaging
CoV replication cycle
Membrane (M) protein: M protein is the most abundant structural protein and defines
the shape of the viral envelope. glycosylated in the Golgi apparatus
Envelope (E) protein: the smallest of the major structural proteins , E is abundantly
expressed inside the infected cell,
Role in the assembly and morphogenesis of virions within the cell.

5. Diagnostic tests
for
Coronavirus disease 2019
(COVID-19)
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6. Diagnostics procedure
How paramedical staff will diagnosis of infection caused by
SARS -CoV -2?
Q. What are the percussion soluble be taken for collection of
sample?
Sample Collection
Ans. Collecting the proper respiratory tract specimen at the right
time from the right anatomic site
How to collect sample?
Within 5 – 6 days of the onset of symptoms when high viral
load s in their upper and lower respiratory tract

7. A nasopharyngeal (NP) swab or an oropharyngeal (OP) swab are
often recommended
OP swabs were used much more frequently in China
But
SARS -CoV -2 RNA was detected only in 32% of OP swabs.
Nasopharyngeal (NP) is Best.
The swab must be inserted deeply into the nasal cavity and should
elicit “tears
Where the swab to be inserted?

8.  Swabs should be kept in place for 10 seconds while twirling the
swab three times
Personal Protective Equipment (PPE )
 Swabs should be placed in viral (universal) transport
medium for rapid transportation to the clinical microbiology
laboratory
Bronchoalveolar lavageOther Sample ????
Sputum- Lower respiratory specimens
Yielded the highest SARS -CoV -2 RNA rate
Virus has been detected in blood and stool,

9. Safety Measures for Specimen Processing
should be done in a class II biological
safety cabinet
Safety cabinet should be in a negative
pressure room within the laboratory

10. Diagnostic tests
for
Coronavirus disease 2019
(COVID-19)
Visit for more vedio and Subscribe @ Hitesh Biochem

11. Methods of Testing
Laboratory
Diagnosis
Real -time
RT -PCR assays
Lateral flow
assays
Enzyme-Linked
Immunosorbent
Assay
RT -PCR) test method: preferred testing method for the
diagnosis of COVID19

12. Processing of respiratory specimen for nucleic acid extraction
before real -time RT -PCR
Real-time reverse-transcription polymerase chain
reaction (RT-PCR)
The specimen should be transferred to
lysis buffer
Lysis buffer = guanidinium -based
inactivating agent as well as a non -
denaturing detergent.

13. Real-time reverse-transcription polymerase chain
reaction (RT-PCR)
RNA extraction from Virus: self -enclosed systems integrating
nucleic acid extraction (GeneXpert®)
Used as a frontline test for COVID-19 as it directly tests for the
presence of the virus RNA.
Sensitive and reliable, capable of producing results in 3-4 hours
Occurs in a closed system to minimize false -positive results

14. Real-time reverse-transcription polymerase chain
reaction (RT-PCR)
 Positive -sense, single -stranded RNA genome that can be used for PCR
assays
 The test kit enzymes copy the RNA to DNA, which is amplified to allow
virus detection by using a PCR machine.
 35 billion copies of viral DNA are made for each viral RNA strand
 Fluorescent markers are typically used to bind to the amplified DNA and
produce light, which can be read by the machine to produce the test result

15. Real-time reverse-transcription polymerase chain
reaction (RT-PCR)
A cycle threshold value (Ct -value) less than 40 is defined as a
positive test
RT-PCR is accepted by scientists and medical staff as a robust and well
documented technique.

16. Lateral Flow / Colloidal Gold
Immunochromatography
‘Antibody tests’ ----- used for pregnancy tests.
Currently used to detect antibodies to disease in a
patient's blood
Tests can detect antibody to virus from patient
blood
Detect the presence of active virus by detecting
virus proteins directly
Require a drop of patient blood
Ability to see if patients are currently infected or
have recovered from COVID-19

17. Lateral Flow / Colloidal Gold Immunochromatography
Label (Enzymes or nanoparticles or fluorescence dyes) coated antibody or aptamer
is immobilized at conjugate pad.
A primary antibody or aptamer against target analyte is immobilized over test line.
A secondary antibody or probe against labeled conjugate antibody/aptamer is
immobilized at control zone.

18. Lateral Flow / Colloidal Gold Immunochromatography
The antibody IgG and IgM lateral flow immunoassay tests are
very simple to read:
Test lines will appear if either of the antibody types are found in the sample. The
appearance of lines for IgG or IgM, or both indicate a positive test

19. Enzyme-Linked Immunosorbent Assay (ELISA)
An Enzyme-Linked Immunosorbent Assay (ELISA) is a common biochemical
technique that can be used to detect antigens or antibodies
ELISAs use enzymes linked to antibodies that can attach to the molecule
that is being tested for and cause a colour change that can be measured by
a specialised machine (ELISA Reader)
Allowing cheap and time effective method for batch testing of large numbers
of patient
The most effective ELISA assays in monitoring prior infection detect
antibodies against SARS-CoV-2.
An ELISA detects antibodies produced in patient blood due to infection
with SARS-CoV-2.

20. Enzyme-Linked Immunosorbent Assay (ELISA)
1. A patient sample of blood or serum
2. The patient antibodies to SARS-CoV-2 stick to the SARS-CoV-2 proteins coated on the
bottom
3. Antibodies with enzymes attached are added and stick to patient antibodies present in
the well
4. Special colourless molecule is added to the well.
Interpretation: An antibody test using ELISA would show a positive result (colour
change) if the patient has antibodies to COVID-19.

21. Drug used for treatment
Chloroquine approved for emergency use by US
FDA
(hydroxychloroquine/Plaquenil, used to treat malaria
and arthritis)
Favilavir, the first approved coronavirus drug in
China
An anti-viral drug, as a treatment for coronavirus.

22. Vaccine under development
University of Oxford’s ChAdOx1 nCoV-19 is an
adenovirus vaccine vector developed by the
university’s Jenner Institute
 Alabama at Birmingham (UAB) to develop a single
dose intranasal vaccine for COVID-19 named
AdCOVID.
DNA vaccine by Entos Pharmaceuticals