Products > 293 Transfection Reagent (Emb Kidney, CRL-1573) Altogen labs Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > 293 Transfection Reagent (Emb Kidney, CRL-1573) Altogen Biosystems offers the 293 Transfection Reagent among a host of 100+ cell line specific In Vitro Transfection Kits. The 293 Transfection Reagent is an advanced formulation of a lipid-based reagent, and it has been developed to provide high transfection efficiency with the 293 cell line. This cell line has applications for studying virucide or efficacy. When cultured in vitro, 293 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 293 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  techserv@altogen.com

293 Cell Line Description and Applications Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > 293 Transfection Reagent (Emb Kidney, CRL-1573) 293 Cell Line Description and Applications The 293 cell line originates from tumorigenic embryonic kidney tissue from a human fetus. The cell line is also known as “Human Embryonic Kidney 293 Cells” or HEK Cells. It is a hypotriploid cell line with a modal chromosome number of 64. HEK 293 Cells grow in culture and transfect efficiently, and as a result, these cells have been a staple in biomedical research for several years. In particular, HEK 293 cells are vital for the development of therapeutic proteins and viruses related to gene therapy. With several transfection techniques, such as the calcium phosphate method, it is possible to achieve 100% transfection with the 293 cell line. High Density 293 cells Atcc.org Altogen Biosystems  848 Rainbow Blvd #823  Las Vegas  NV  89107  USA Telephone  702 349 6103  Fax  702 989-0841  email  techserv@altogen.com

293 Transfection Protocol Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > 293 Transfection Reagent (Emb Kidney, CRL-1573) 293 Transfection Protocol 1. Plate 10,000 - 15,000 293 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 293 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 293 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  techserv@altogen.com

General Lipoplex-mediated Transfection Mechanism of Action Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > 293 Transfection Reagent (Emb Kidney, CRL-1573) General Lipoplex-mediated Transfection Mechanism of Action Altogen Biosystems  848 Rainbow Blvd #823  Las Vegas  NV  89107  USA Telephone  702 349 6103  Fax  702 989-0841  email  techserv@altogen.com

293 Transfection Kit Product Details Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > 293 Transfection Reagent (Emb Kidney, CRL-1573) 293 Transfection Kit Product Details Two-component formulation enhances lipid-mediated transfection efficiency Optimized for intracellular delivery of plasmid DNA, siRNA, microRNA, and mRNA High transfection efficiency of both siRNA and plasmid DNA without compromising cell viability Achieve robust siRNA uptake for dependable gene silencing Effective transfection under conditions of up to 40% serum Transfection kit includes Transfection Enhancer reagent Gentle enough to be used for single cell analysis 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  techserv@altogen.com

Data Products > 293 Transfection Reagent (Emb Kidney, CRL-1573) Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > 293 Transfection Reagent (Emb Kidney, CRL-1573) Data Figure 1. GAPD mRNA levels were quantified using real-time RT-PCR in the 293 cells transfected with siRNAs targeting GAPD or non-silencing siRNA. Forty-eight hours post-transfection, the cells were harvested and analyzed by real-time RT-PCR for GAPD mRNA expression levels. Data were normalized against the 18S rRNA signal. Control samples were either mock-transfected or untreated. Values are normalized to untreated sample. Data are means ± SD (n=5).

Data Products > 293 Transfection Reagent (Emb Kidney, CRL-1573) Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > 293 Transfection Reagent (Emb Kidney, CRL-1573) Data Figure 2. Protein expression of GAPDH in 293 cells. DNA plasmid expressing GAPDH or siRNA targeting GAPDH were transfected into 293 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.

293 Transfection Kit Benefits Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > 293 Transfection Reagent (Emb Kidney, CRL-1573) 293 Transfection Kit Benefits Pre-optimized transfection protocol for 293 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 293 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 293 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  techserv@altogen.com