Stratagene PathDetect Reporter Cell Lines INSTRUCTION MANUAL
PathDetect Reporter Cell Lines
INSTRUCTION MANUAL
Catalog #800050 (PathDetect HLR Cell Line)
#800055 (HLR-Elk1 Cell Line)
#800060 (HLR-CHOP Cell Line)
#800065 (HLR-CREB Cell Line)
Revision A
BN #800050-12
For In Vitro Use Only
800050-12
LIMITED PRODUCT WARRANTY
This warranty limits our liability to replacement of this product. No other warranties of any kind,
express or implied, including without limitation, implied warranties of merchantability or fitness for
a particular purpose, are provided by Agilent. Agilent shall have no liability for any direct, indirect,
consequential, or incidental damages arising out of the use, the results of use, or the inability to use
this product.
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PathDetect Reporter Cell Lines
CONTENTS
Materials Provided.............................................................................................................................. 1
Storage Conditions.............................................................................................................................. 1
Additional Materials Required .......................................................................................................... 2
Safety Consideration........................................................................................................................... 2
Notices to Purchaser ........................................................................................................................... 2
Introduction......................................................................................................................................... 3
Preprotocol Considerations................................................................................................................ 5
Choosing a Transfection Method .......................................................................................... 5
Tissue Cultureware................................................................................................................ 5
Designing the Experiment ..................................................................................................... 5
Cell Culture Protocols......................................................................................................................... 7
Starting the Cells in Culture .................................................................................................. 7
Passaging the Cells................................................................................................................ 7
Freezing the Cells for Long-term Storage ............................................................................. 8
Preparing the Cells for Transfection................................................................................................. 9
Preparing Plasmids for Transfection ................................................................................................9
Studying the Effects of a Gene Product................................................................................. 9
Transfecting the Cells ......................................................................................................................... 9
Preparing Cell Lysates for Luciferase Activity Assay ................................................................... 10
Performing the Luciferase Activity Assay ...................................................................................... 10
Troubleshooting ................................................................................................................................ 11
Appendix I: Plasmid Information.................................................................................................... 12
The pFA trans-Activator Plasmids...................................................................................... 12
Preparation of Media and Reagents................................................................................................ 13
References .......................................................................................................................................... 14
Endnotes............................................................................................................................................. 14
MSDS Information............................................................................................................................ 14
Quick-Reference Protocol ................................................................................................................ 15
PathDetect Reporter Cell Lines
ATERIALS PROVIDED
M
Catalog
#800050
HLR Cell Line — 1 × 106 viable
pFA2-CREB fusion trans-activator plasmid 25 ng/μl 5 μg
pFC-PKA plasmid (positive control) 25 ng/μl 5 μg
pFC-CMV negative control plasmid 25 ng/μl 5 μg
Catalog
#800055
HLR-Elk1 trans-Reporting Cell Line — 1 × 106 viable
pFC-CMV negative control plasmid 25 ng/μl 5 μg
pFC-MEK1 plasmid (positive control) 25 ng/μl 5 μg
Catalog
#800060
HLR-CHOP trans-Reporting Cell Line — 1 × 106 viable
pFC-CMV negative control plasmid 25 ng/μl 5 μg
pFC-MEK3 plasmid (positive control) 25 ng/μl 5 μg
Catalog
#800065
HLR-CREB trans-Reporting Cell Line — 1 × 106 viable
pFC-CMV negative control plasmid 25 ng/μl 5 μg
pFC-PKA plasmid (positive control) 25 ng/μl 5 μg
Component Concentration Quantity
cells
Component Concentration Quantity
cells
Component Concentration Quantity
cells
Component Concentration Quantity
cells
STORAGE CONDITIONS
Control Plasmids: –20°C
Cell lines: Liquid Nitrogen
Revision A © Agilent Technologies, Inc. 2008.
PathDetect Reporter Cell Lines 1
ADDITIONAL MATERIALS REQUIRED
Growth medium (See Preparation of Media and Reagents)
Luciferase assay kit
5-ml BD Falcon polystyrene round-bottom tubes (BD Biosciences Catalog #352054)
Calcium- and magnesium-free PBS
Tissue culture plasticware
Transfection reagents
Luminometer
Hygromycin
G418 Sulfate
SAFETY CONSIDERATION
The American Type Culture Collection (ATCC) has designated CCL-2 HeLa cell lines as Biosafety
Level 2. Appropriate caution should be used while handling any human-derived cell line. For more
information, see Biosafety in Microbiological and Biomedical Laboratories (4th edition). It is
available on the Web sites of both the National Institutes of Health at http://bmbl.od.nih.gov and the
Office of Health and Safety of the Centers for Disease Control and Prevention at
www.cdc.gov/od/ohs (Click the link to Biosafety Information).
NOTICES TO PURCHASER
The use of the CMV Promoter is covered under U.S. Patent Nos. 5,168,062 and 5,385,839 owned by
the University of Iowa Research Foundation and licensed FOR RESEARCH USE ONLY. For
further information, please contact UIRF at 319-335-4546.
A license (from Promega for research reagent products and from The Regents of the University of
California for all other fields) is needed for any commercial sale of nucleic acid contained within or
derived from this product.
The HLR, HLR-cJun, HLR-Elk1, HLR-CHOP, and HLR-CREB cell lines are proprietary to Agilent.
Purchase of these cell lines includes a limited non-exclusive license under Agilent’s proprietary
rights to use these cell lines for research purposes only. This license does not grant any rights to: (i)
modify any of these cell lines; (ii) offer any of these cell lines, or any derivatives or modifications
thereof, for resale; or (iii) otherwise distribute or transfer any of these cell lines, or any derivative or
modification thereof, to third parties for any purpose or use. No other license, express, implied or by
estoppel, is granted.
2 PathDetect Reporter Cell Lines
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