Transcript PPT - Altogen Biosystems

Leading Developer and Manufacturer of In Vivo and DNA Transfection
Kits, Transfection Reagents and Electroporation Delivery Products
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Keratinocyte Transfection Reagent (Keratinocytes)
Altogen Biosystems offers the Keratinocyte Transfection Reagent
among a host of 100+ cell line specific In Vitro Transfection Kits.
Keratinocyte Transfection Reagent is an advanced cationic lipid
formulation based reagent, and it has been developed to provide
high transfection efficiency with Keratinocyte cells.
This cell line is a good host for studying skin and research areas
related to keratinocytes. When cultured in vitro, Keratinocyte cells
are a monolayer and adhere to the surface of the culture flask.
Other applications include drug discovery, gene expression
studies, molecular and cell biology research applications.
Purchase Keratinocyte Transfection Kit at www.Altogen.com
Altogen Biosystems  848 Rainbow Blvd #823  Las Vegas  NV  89107  USA
Telephone  702 349 6103  Fax  702 989-0841  email  [email protected]
Leading Developer and Manufacturer of In Vivo and DNA Transfection
Kits, Transfection Reagents and Electroporation Delivery Products
Keratinocyte Transfection Reagent (Keratinocytes)
Keratinocyte Cell Line Description and Applications
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Keratinocyte Cells
www.sciencellonline.com
Keratinocyte cells are the most common type
of skin cell; they form in the basal skin layer
and grow through epidermis layers.
Keratinocytes produce keratin and are also the
primary source of proinflammatory cytokine IL1. These cells are crucial to skin production
and make up approximately 95% of the
epidermis cells. Keratinocyte cells weave
between nerves and epidermis tissue and
provide protection from potentially infectious
substances. Keratinocytes can be studied in
vitro to assess skin irritants.
Altogen Biosystems  848 Rainbow Blvd #823  Las Vegas  NV  89107  USA
Telephone  702 349 6103  Fax  702 989-0841  email  [email protected]
Leading Developer and Manufacturer of In Vivo and DNA Transfection
Kits, Transfection Reagents and Electroporation Delivery Products
Keratinocyte Transfection Reagent (Keratinocytes)
Keratinocyte Transfection Protocol
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1. Plate 10,000 - 15,000 Keratinocyte cells per well in 0.5 ml of complete growth medium 12–24 hours prior
to transfection
2. Wash with 1xPBS and add 0.5 ml of fresh growth medium
3. Prepare transfection complexes by mixing 40 µl of serum-free medium, 5.5 µl of transfection reagent,
and
• 750 ng DNA (or mRNA), or
• 30 nM - 50 nM of siRNA (or microRNA)
*Referred to a final volume including growth medium
4. Incubate transfection complexes at RT for 15 - 30 minutes
5. Optional: Add 2 µl of Complex Condenser. This reagent reduces the size of transfection complex,
therefore increasing transfection efficiency; however it may increase cell toxicity
6. Add prepared transfection complexes to 0.5 ml of complete growth medium with Keratinocyte cells
(from step 2)
7. Incubate cells at 37ºC in a humidified CO2 incubator
8. Assay for phenotype or target gene expression 48 - 72 hours after transfection
Optional: Transfection efficiency can be increased by addition of Transfection Enhancer reagent. Add 2 µl of
Transfection Enhancer reagent 12-24 hours after transfection
If the viability of Keratinocyte cells being transfected is affected at 16 - 24 hours post-transfection, the level of
cytotoxicity can be decreased by changing the growth medium and eliminating redundant exposure of cells to
transfectant
Altogen Biosystems  848 Rainbow Blvd #823  Las Vegas  NV  89107  USA
Telephone  702 349 6103  Fax  702 989-0841  email  [email protected]
Leading Developer and Manufacturer of In Vivo and DNA Transfection
Kits, Transfection Reagents and Electroporation Delivery Products
Keratinocyte Transfection Reagent (Keratinocytes)
General Lipoplex-mediated Transfection Mechanism of Action
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Altogen Biosystems  848 Rainbow Blvd #823  Las Vegas  NV  89107  USA
Telephone  702 349 6103  Fax  702 989-0841  email  [email protected]
Leading Developer and Manufacturer of In Vivo and DNA Transfection
Kits, Transfection Reagents and Electroporation Delivery Products
Keratinocyte Transfection Reagent (Keratinocytes)
Keratinocyte Transfection Kit Product Details
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•
Two component formulation enhances lipid mediated transfection efficiency
•
Optimized easy-to-use transfection protocol provided for transfection of siRNA, DNA,
mRNA, and microRNA
•
Kit includes Transfection Enhancer reagent and recommended transfection protocol
•
High transfection efficacy in the presence of serum
•
Expand your RNAi application with a reagent optimized for delivery of both siRNA and
plasmid
•
Reproducible transfection results
•
Works well for standard reverse transfection and high-throughput applications
Developed and manufactured by Altogen Biosystems (www.Altogen.com)
Altogen Biosystems  848 Rainbow Blvd #823  Las Vegas  NV  89107  USA
Telephone  702 349 6103  Fax  702 989-0841  email  [email protected]
Leading Developer and Manufacturer of In Vivo and DNA Transfection
Kits, Transfection Reagents and Electroporation Delivery Products
Products >
Keratinocyte Transfection Reagent (Keratinocytes)
Data
Figure 1. Cyclophilin B silencing efficiency was determined by qRT-PCR in the cells transfected by
Cyclophilin B siRNA or non-silencing siRNA control following the recommended transfection protocol.
Cyclophilin mRNA expression levels were measured 48 hours post-transfection. 18S rRNA levels were used
to normalize the Cyclophilin B data. Values are normalized to untreated sample. Data are presented as
means ± SD (n=3).
Leading Developer and Manufacturer of In Vivo and DNA Transfection
Kits, Transfection Reagents and Electroporation Delivery Products
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Keratinocyte Transfection Reagent (Keratinocytes)
Data
Figure 2. Protein expression of Cyclophilin B in Keratinocyte cells. DNA plasmid expressing Cyclophilin B or
siRNA targeting Cyclophilin B were transfected into Keratinocyte cells following Altogen Biosystems transfection
protocol. At 72 hours post-transfection the cells were analyzed by Western Blot for protein expression levels
(normalized by total protein, 10 µg of total protein loaded per each well). Untreated cells used as a negative
control.
Leading Developer and Manufacturer of In Vivo and DNA Transfection
Kits, Transfection Reagents and Electroporation Delivery Products
Keratinocyte Transfection Reagent (Keratinocytes)
Keratinocyte Transfection Kit Benefits
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• Pre-optimized transfection protocol for Keratinocyte cell line
• Compatible with DNA, small RNA (siRNA, shRNA, miRNA), mRNA, and small protein complexing
• Free of serum and protein of animal origin
• Compatible with standard and reverse transfection methods (both protocols provided in the kit manual)
• Easy to use Keratinocyte transfection protocol ensures great performance with expedited experimental
timeline
• Equally efficient for single or multiple transfections
• Can be used for transient transfection and development of stable Keratinocyte cell lines
• Bio-degradable after endocytosis
• Used for preclinical research worldwide
Altogen Biosystems  848 Rainbow Blvd #823  Las Vegas  NV  89107  USA
Telephone  702 349 6103  Fax  702 989-0841  email  [email protected]