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High
Transfection Efficiency
High
Co-Transfection Efficiency
PCR Products Can Be
Used Directly
Flexible Assay Type
and Format
Avoid Problems
with Stable Cell Lines
High Transfection Efficiency
STEP routinely achieves significantly
higher transfection efficiency than convential
liquid-based transfection methods.
HEK293T:
Neuro-2A:
SH-SY5Y:
The STEP procedure has been tested on more
than 30 cell lines and the following is the transfection effciency
for some of the popular cell lines:
HEK
293 cells 80%
HEK
293T cells >90%
PC12
cells >30%
Neuro-2A
>70%
SH-SY5Y
neuroblastomas >50%
SHEP
neuroblastomas 40-50%
C6
glioma cells 40-50%
HepG2
liver cells 40%
BC3H1
smooth muscle cells 40%
High Co-Transfection Efficiency
STEP can approach 100% co-transfection
efficiency on cell lines with high transfection rate. With cell lines that
are difficult to transfect using convention approach, STEP still exhibits
superior transfection and co-transfection efficiency. The following pictures
were taken from SH-SY5Y neuronal cells transfected
with a DNA mixture of 5% pZsGreen1-C1, 5% pAsRed2-C1, and 90% pTAL-SEAP (as control DNA). The overall transfection
efficiency was about 40%. Around 90% transfected
cells co-expressed the red and green fluorescence proteins.
Such high
co-transfection efficiency allows fine tuning the expression of each
component required in a reporter assay. In addition, multiple reporter assays
can easily be co-introduced into a cell.
PCR Products Can Be Used Directly
Most transient transfection methods require
supercoiled plasmids for good transfection
efficiency. Constructing and preparing plasmids can be time-consuming for
large-scale genomic studies. STEP can express PCR products containg a CMV promoter-driven coding region efficiently.
Flexible
Assay Type and Format
STEP ssays can
use endogenous cellular responses or transfected
reporter molecules including:
Individual response elements
Complex promoters
Enzymes and other proteins
Any effector
plasmid can be used in STEP-based assays:
Signaling proteins
Transcription factors
Metabolic enzymes
Transporters
STEP results can be monitored by a number
of measures including:
Fluorescence
FLIPR
Fluorescent proteins (GFP, DsRed, calcein)
ß-Lactamase (GeneBlazer
TM)
Luminescence
Secreted Alkaline Phosphatase (SEAP)
Luciferase
Colorimetric
Microscopic
Cell morphology
Subcellular compartmentation
Cell type and incubation conditions can be
modified to achieve optimal assay requirements
STEP has been implemented in 96-well plate,
384-well plate as well as the microarray format on
microscope slides. Microarrays of STEP complexes on
multi-well plate is being developed to further increase the throughput of
screening and enrich the information obtained from each single well.
Avoid Problems with Stable Cell Lines
STEP offers highly efficient and consistent
transient transfections for high throughput drug
screening and functional genomic studies. It obsoletes the need for making
and using stable cell lines for most applications. In addition to much faster
assay setup and cost reduction, it also enable the
functional assay of toxic genes due to its transient nature. Furthermore, the
high co-transfection effciency of STEP method will
allow the study of multi-subunit proteins or complex signal transduction
pathways.
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