MOLECULAR DIAGNOSTIC

Dyke Gita Wirasisya, S.Farm., M.Sc., Apt.

Program Studi Farmasi
Universitas Mataram
What is pathogens? And how to detect it?
• Direct culture

• Biochemical assay

• Immunological assay

• Nucleic acid test

What is the structure and function of DNA?
How DNA replicate?
What is the function of PCR?
What is Primer?
INFECTIOUS PATHOGENS
HIV and TB
Case of HIV Detection
Diagnostic HIV

• Symptomps are non

specific.

• Detect antigen,
antibodies, spesific
protein or RNA
Window periode of HIV Infection
ELISA (Enzime-Linked Immunosorbent Assay)
 Lysing agend Test band Control band
Bound Labled AB. (bound AB) (bound AB)
AB

Free labled
AB

Nitrocellulose strip

http://www.rapid-diagnostics.org/

Dye-labelled antibody, specific for target antigen, is present on the

lower end of nitrocellulose strip or in a plastic well provided with the
strip

Antibody, also specific for the target antigen, is bound to the strip in
a thin (test) line, and either antibody specific for the labelled
antibody, or antigen, is bound at the control line
Source: http://www.wpro.who.int/rdt
Laboratory Training for Field Epidemiologists
 Immunochromatography
Parasitized
Blood
Test band Control band
(bound AB) (bound AB)

Parasite antigen (AG.)

Captured by labled AB.

Blood and labled Ab flushed along the strip

Blood and buffer, which have been placed on strip or in the well,
are mixed with labelled antibody and are drawn up strip across
the lines of bound antibody
Source: http://www.wpro.who.int/rdt
Immunochromatography

Captured Ag-labled Captured labled Ab

Ab-complex

Labled AB-AG- Labled AB-AG-

complex complex
Captured by Captured by
bound AB of bound AB of
test band control band

If antigen is present, some labelled

antibody will be trapped on the test line.
Excess-labelled antibody is trapped on
the control line
Source: http://www.wpro.who.int/rdt
HIV
• HIV 1 • HIV 2
 HIV-1 and HIV-2 Western blot result…

NAT is used to differentiate

HIV-1 and HIV-2 fastly…
Case of MTB Detection

• Conventional detection
using sputum and
staining…
• For sensitivity test
need 4-8 weeks!
• Resistent to rifampicin in MTB caused by ropB mutation.
• Conventional PCR only detect the gene of MTB but
cannot detect whether MTB resistent to rifampicin or not.
• To determine it by using RT-Q-PCR (Real Time-
quantitative-PCR) using 5 different molecular beacon.

Molecular Beacons
rpoB gene and molecular beacons
TERIMAKASIH