1. Products > VERO Transfection Reagent (Monkey Kidney Cells) Altogen Biosystems offers the VERO Cell Transfection Reagent among a host of 100+ cell line specific In Vitro Transfection Kits. The VERO Cell Transfection Reagent is a lipid mediated formulation, and it has been developed to provide high transfection efficiency with VERO. This cell line is a good host for studying a range of areas within the field of biomedical therapeutics and biology including toxicology and virology. When cultured in vitro, VERO 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 VERO Transfection Kit at www.Altogen.com Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Altogen Biosystems  848 Rainbow Blvd #823  Las Vegas  NV  89107  USA Telephone  702 349 6103  Fax  702 989-0841  email  techserv@altogen.com

2. VERO Cell Line Description and Applications Altogen Biosystems  848 Rainbow Blvd #823  Las Vegas  NV  89107  USA Telephone  702 349 6103  Fax  702 989-0841  email  techserv@altogen.com High Density VERO cells atcc.org Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > VERO Transfection Reagent (Monkey Kidney Cells) The Vero cell line originates from the kidney epithelial cells taken from an African green monkey (Cercopithecus aethiops). Yasumura and Kawakita established the cell line in 1962 and named the cell line as an abbreviation of “verda reno” meaning “green kidney” in Esperanto. Vero cells are aneuploidy, continuous, and can be replicated through several division cycles. Unique from mammalian cells, Vero cells do not produce type 1 interferons post infection. The Vero cell line has several applications ranging from serving as host for the growing viruses and eukaryotic parasites (trypanosomatids in particular.) The Vero cell line can also be used be using for the screening of toxins such as Escherichia coli.

3. VERO Transfection Protocol 1. Plate 10,000 - 15,000 VERO 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 VERO cells (from step 2) 7. Incubate cells at 37ºC in a humidified CO 2 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 VERO 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 Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > VERO Transfection Reagent (Monkey Kidney Cells)

4. 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 Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > VERO Transfection Reagent (Monkey Kidney Cells)

5. VERO Transfection Kit Product Details 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  techserv@altogen.com Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > VERO Transfection Reagent (Monkey Kidney Cells)

6. Data Figure 1. GAPD mRNA levels were quantified using real-time RT-PCR in the VERO 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 GAPDH 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=3). Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > VERO Transfection Reagent (Monkey Kidney Cells)

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

8. Pre-optimized transfection protocol for VERO 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 VERO 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 VERO cell lines Bio-degradable after endocytosis Used for preclinical research worldwide VERO Transfection Kit Benefits Altogen Biosystems  848 Rainbow Blvd #823  Las Vegas  NV  89107  USA Telephone  702 349 6103  Fax  702 989-0841  email  techserv@altogen.com Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > VERO Transfection Reagent (Monkey Kidney Cells)