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- : S7750
- : 96 assays
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Clinical research samples (1-10) and dilutions of telomerase positive cells were subjected to the TRAPeze® ELISA Telomerase Detection Kit protocol. At...
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TRAPeze® ELISA Telomerase Detection Kit, strip well plates
Description:
TRAPeze® ELISA Telomerase Detection Kit, strip well plates
Trade Name:
Chemicon (Millipore)
Qty/Pk:
96 assays
Product Overview:
The TRAPeze® ELISA Telomerase Detection Kit is a rapid, non-isotopic assay for detecting telomerase activity in cell or tissue extracts. The method combines a one-step TRAP assay (PCR) with direct, chromogenic detection of TRAP products (ELISA).
Recommended Taq polymerases: must be non-proofreading, having no exonuclease activity and capable of "hot-start." Titanium Taq™, Platinum® Taq are suggested. The kit uses coated strip-well pieces for ease of use and handling.
Using this Manual
The TRAPEZE® ELISA Telomerase Detection Kit provides reagents necessary to perform the Telomeric Repeat Amplification Protocol (TRAP) and ELISA assay for non-quantitative detection of telomerase activity in cells and tissues.
The novice user is advised to read the entire manual prior to using the TRAPEZE® ELISA Kit, particularly Sec. III: Protocol. Supplemental protocols can be found in Sec. V. Appendix. Should additional questions arise, assistance can be obtained by contacting Chemicon Technical Support at (800) 437-7500 or at techserv@chemicon.com.
Recommended Taq polymerases: must be non-proofreading, having no exonuclease activity and capable of "hot-start." Titanium Taq™, Platinum® Taq are suggested. The kit uses coated strip-well pieces for ease of use and handling.
Using this Manual
The TRAPEZE® ELISA Telomerase Detection Kit provides reagents necessary to perform the Telomeric Repeat Amplification Protocol (TRAP) and ELISA assay for non-quantitative detection of telomerase activity in cells and tissues.
The novice user is advised to read the entire manual prior to using the TRAPEZE® ELISA Kit, particularly Sec. III: Protocol. Supplemental protocols can be found in Sec. V. Appendix. Should additional questions arise, assistance can be obtained by contacting Chemicon Technical Support at (800) 437-7500 or at techserv@chemicon.com.
Background Information:
Telomeres are specific structures found at the end of chromosomes in eukaryotes. In human chromosomes, the telomeres consist of thousands of copies of 6 base repeats (TTAGGG)(1-3). It has been suggested that telomeres protect chromosome ends since damaged chromosomes lacking telomeres undergo fusion, rearrangement and translocation (2). In somatic cells, telomere length is progressively shortened with each cell division both in vivo and in vitro (4-7) due to the inability of the DNA polymerase complex to synthesize the very 5' end of the lagging strand (8,9).
Telomerase is a ribonucleoprotein that synthesizes and directs the telomeric repeats onto the 3' end of existing telomeres using its RNA component as a template (10-14). Telomerase activity has been shown to be specifically expressed in immortal cells, cancer and germ cells (15,16) where it compensates for telomere shortening during DNA replication and thus stabilizes telomere length (7,17). These observations have led to a hypothesis that telomere length may function as a "mitotic clock" to sense the number of cell divisions and eventually signal replicative senescence or programmed cell death when a critical telomere length is achieved. Therefore, expression of telomerase activity in cancer cells may be a necessary and essential step for tumor development and progression (16,18-20). The causal relationship between expression of telomerase and telomere length stabilization and the extension of the life span of the human cell has recently been reported (21).
The development of a sensitive and efficient PCR-based telomerase activity detection method, TRAP (Telomeric Repeat Amplification Protocol)(15, 22), has made possible large scale surveys of telomerase activity in human cells and tissues (15, 23-29). To date, telomerase activity has been detected in over 85% of all tumors tested spanning more than 20 different types of cancers (30-31).
The TRAPEZE® ELISA Kit provides the reagents necessary for performing the TRAP assay followed by ELISA detection of telomerase activity in cell/tissue samples.
Telomerase is a ribonucleoprotein that synthesizes and directs the telomeric repeats onto the 3' end of existing telomeres using its RNA component as a template (10-14). Telomerase activity has been shown to be specifically expressed in immortal cells, cancer and germ cells (15,16) where it compensates for telomere shortening during DNA replication and thus stabilizes telomere length (7,17). These observations have led to a hypothesis that telomere length may function as a "mitotic clock" to sense the number of cell divisions and eventually signal replicative senescence or programmed cell death when a critical telomere length is achieved. Therefore, expression of telomerase activity in cancer cells may be a necessary and essential step for tumor development and progression (16,18-20). The causal relationship between expression of telomerase and telomere length stabilization and the extension of the life span of the human cell has recently been reported (21).
The development of a sensitive and efficient PCR-based telomerase activity detection method, TRAP (Telomeric Repeat Amplification Protocol)(15, 22), has made possible large scale surveys of telomerase activity in human cells and tissues (15, 23-29). To date, telomerase activity has been detected in over 85% of all tumors tested spanning more than 20 different types of cancers (30-31).
The TRAPEZE® ELISA Kit provides the reagents necessary for performing the TRAP assay followed by ELISA detection of telomerase activity in cell/tissue samples.
Key Applications:
PCR
Species Reactivity:
Mammals
Usage Statement:
Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
Components:
- 1. CHAPS XL Lysis Buffer (10 mL)
-
2. 5X TRAPEZE® Reaction Mix (1 mL)
• biotinylated TS primer
• biotinylated RP primer
•dA, dC, dG and dTTP
•internal control primers
•Tris buffer -
3. PCR - Grade Water (10 mL)
protease, DNase, and RNase-free;
deionized -
4. TSR8* (control template) (32 µL)
0.1 amole/µL TSR8 template -
5. Control Cell Pellet
Telomerase positive cells (106 cells) -
6. Anti-DNP Antibody, HRP-conjugated
(22µL) - 7. Blocking/Dilution Buffer (45 mL)
- 8. 10X Washing Buffer (45 mL)
-
9. TMB Solution (10 mL)
Tetramethylbenzidine (HRP substrate) - 10. Stop Solution (10 mL)
- 11. Streptavidin-coated Microtiter Plate
- * Caution - refer to Sec. II. Kit Components, Warning and Precautions.
Brand Family:
Chemicon
Storage Conditions:
Precautions
1. Because the TRAPEZE® ELISA Telomerase Detection Kit detects the activity of telomerase, a RNase sensitive ribonucleoprotein, and not merely the presence of the RNA or protein components of telomerase, the assay requires enzymatically active cell or tissue samples. Furthermore, due to the sensitivity of the TRAP assay, which can detect telomerase activity in a very small number of cells, a special laboratory setup and significant precautions are required to prevent PCR carry-over contamination and RNase contamination. These precautions are discussed in detail in Sec. V. Appendix, Laboratory Setup and Precautions and TRAP Station Setup.
Before performing a TRAP assay, it is strongly recommended that you read the entire manual. At a minimum, read Sec. V. Appendix, Laboratory Setup and Precautions and Sec. III. Protocol, Experimental Design which describe all the controls that must be included in each analysis.
2. For Research Use Only. Not for use in diagnostic procedures.
1. Because the TRAPEZE® ELISA Telomerase Detection Kit detects the activity of telomerase, a RNase sensitive ribonucleoprotein, and not merely the presence of the RNA or protein components of telomerase, the assay requires enzymatically active cell or tissue samples. Furthermore, due to the sensitivity of the TRAP assay, which can detect telomerase activity in a very small number of cells, a special laboratory setup and significant precautions are required to prevent PCR carry-over contamination and RNase contamination. These precautions are discussed in detail in Sec. V. Appendix, Laboratory Setup and Precautions and TRAP Station Setup.
Before performing a TRAP assay, it is strongly recommended that you read the entire manual. At a minimum, read Sec. V. Appendix, Laboratory Setup and Precautions and Sec. III. Protocol, Experimental Design which describe all the controls that must be included in each analysis.
2. For Research Use Only. Not for use in diagnostic procedures.
Packaging:
Strip-well plate
Product Name:
TRAPeze® ELISA Telomerase Detection Kit, strip well plates
Materials Required but Not Delivered:
Equipment and Supplies
1. Thermocycler
2. ELISA plate reader (neeed 450 nm and 690 nm filters)
5. If analyzing tissues, homogenization equipment as described in Sec. III. Protocol, Extract Preparation
6. Tubes for PCR amplification
7. Aerosol resistant pipette tips (RNase-free)
8. Laboratory film
Reagents
1. Taq polymerase (cloned, unmodified)
2. PBS (Mg2+- and Ca2+-free)
3. Reagents for protein concentration measurement (See Sec. V. Appendix, Determination of Protein Concentration)
4. RNase inhibitor (for extract preparation from tissues)
5. Buffer used with the analytical spectrofluorometer (Option 1) (See Sec. III. Protocol, TRAPEZE® ELISA Telomerase Detection Kit Assay)
1. Thermocycler
2. ELISA plate reader (neeed 450 nm and 690 nm filters)
5. If analyzing tissues, homogenization equipment as described in Sec. III. Protocol, Extract Preparation
6. Tubes for PCR amplification
7. Aerosol resistant pipette tips (RNase-free)
8. Laboratory film
Reagents
1. Taq polymerase (cloned, unmodified)
2. PBS (Mg2+- and Ca2+-free)
3. Reagents for protein concentration measurement (See Sec. V. Appendix, Determination of Protein Concentration)
4. RNase inhibitor (for extract preparation from tissues)
5. Buffer used with the analytical spectrofluorometer (Option 1) (See Sec. III. Protocol, TRAPEZE® ELISA Telomerase Detection Kit Assay)