In: Biology
Could you explain to me how TRAP assay works? I want to detect telomerase activity of breast cancer cell after the cell treated with anti-tumor (curcumin).
The principle of TRAP assay
The TRAP assay is based on telomerase activity. Telomeres are
present at the linear end of chromosomes and generally surrounded
by proteins. TRAP assay is used to measure the telomerase activity
in mammalian cells and tissue samples. It basically consists of
three steps extension, amplification, and detection of telomerase
activity.
Extension
step: In this step telomeric repeats are added
to the telomerase substrate.
Amplification:
In the amplification step the telomerase product is amplified by
Polymerase chain reaction (PCR) using specific upstream and
downstream primers.
Detection
phase: The presence or absence of telomerase
is confirmed by analyzing gel electrophoresis. To avoid false
positive results an internal standard control is used TSNT which is
amplified by TS primers.
This principle can be extrapolated to study the telomerase activity
of breast cancer after treating the cell with curcumin.