1. Products > PC-3 Transfection Reagent (Prostate Cancer Cells) Altogen Biosystems offers the PC-3 Transfection Reagent among a host of 100+ cell line specific In Vitro Transfection Kits. The PC-3 Transfection Reagent is an advanced formulation of a lipid based reagent, and it has been developed to provide high transfection efficiency with the PC-3 cell line. This cell line is considered to be a classic cell line for studying prostate cancer. When cultured in vitro, PC-3 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 PC-3 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. PC-3 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 PC-3 cells Atcc.org Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > PC-3 Transfection Reagent (Prostate Cancer Cells) The PC-3 cell line originates from the bone marrow metastasis of a 62-year-old Caucasian man with grade IV prostate cancer post mortem. The patient was treated with androgen suppression therapy. This cell line has a high metastatic potential and research indicates that the cells will form tumors in nude mice. The PC-3 cell line expresses Prostate Specific Antigen and has a low amount of testosterone-5-alpha reductase activity. This cell line has unique applications for research related to cancer and molecular and cellular biology due to its high metastatic potential. The PC-3 cell line is also considered to be a classic cell line in the study of prostate cancer.

3. PC-3 Transfection Protocol 1. Plate 10,000 - 15,000 PC-3 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 PC-3 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 PC-3 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 > PC-3 Transfection Reagent (Prostate Cancer 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 > PC-3 Transfection Reagent (Prostate Cancer Cells)

5. PC-3 Transfection Kit Product Details Cationic lipids formulation 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 Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > PC-3 Transfection Reagent (Prostate Cancer Cells)

6. Data Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Figure 1. GAPD mRNA levels were quantified using real-time RT-PCR in the PC3 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). Products > PC-3 Transfection Reagent (Prostate Cancer 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 PC-3 cells. DNA plasmid expressing GAPDH or siRNA targeting GAPDH were transfected into PC-3 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 > PC-3 Transfection Reagent (Prostate Cancer Cells)

8. Pre-optimized transfection protocol for PC-3 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 PC-3 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 PC-3 cell lines Bio-degradable after endocytosis Used for preclinical research worldwide PC-3 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 > PC-3 Transfection Reagent (Prostate Cancer Cells)