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Biomarker Discovery Protocols: This database contains a comprehensive list of various bioprotocols that are used in biomarker discovery. These complete protocols include techniques used in molecular and protein biomarker discovery research. We have listed unique protocols that are not available in any other online bioprotocol database.
Molecular Biomarker Discovery Protocols

Telomeric Repeat Amplification Protocol (TRAP) assay

Telomerase activity from cell extracts was analyzed using a previously described realtime PCR-based telomeric repeat amplification protocol (TRAP) (Xu T, Xu Y, Liao CP, Lau R, Goldkorn A. Reprogramming murine telomerase rapidly inhibits the growth of mouse cancer cells in vitro and in vivo. MoI Cancer Ther 2010;9:438-49). The microfilter with captured cells was lysed in TRAPeze® IX CHAPS Lysis Buffer (Millipore, 5 Temecula, CA), and 5 micro liter of cell lysate per reaction was added for each sample. For each reaction, DU 145 cell lysates were used as standard controls in parallel.

QPCR telomerase activity assay was carried out as described below.

1. TS oligonuleotides (5' AATCCGTCGAGCAGAGTT, 8ng/micro liter) and 50micro molar of each dNTPs are added to the cell lysate and incubated on a Bio-Rad MyiQ Thermocycler system for 30 min at 30 °C followed by inactivation of telomerase at 95 0C for 1 min.2. ACX reverse primer (5'GCGCGGCTTACCCTTACCCTTACCCTAACC, 4ng/micro liter), SYBR green (1 :20,000) and 0.8U Taq platinum polymerase (Invitrogen, Carlsbad, CA) are added to the same tube in a master mix formula.

The reaction is run on the same machine for 40 cycles at 95 0C, 0"; 50 0C, 5"; 72 0C, 10", and the iQ5 optical system software version 2.0 was used to analyze the results.

Source: WO/2010/135603

Disclaimer: Sciclips does not guarantee the authenticity or validity of the protocols listed in this section. These protocols are extracted "as is" from various sources, with minor modifications. Please refer to the respective citations, which are hyperlinked, for more details. The use of cited molecules, not limited to proteins, nucleic acids, compounds etc, in the protocols that are extracted from patents may be covered under patent rights. These molecules should be used for information purpose only.
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