Products > IRR-MRC5 Transfection Reagent (Irradiated Fibroblasts) Altogen labs Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > IRR-MRC5 Transfection Reagent (Irradiated Fibroblasts) Altogen Biosystems offers the IRR-MRC5 Transfection Reagent among a host of 100+ cell line specific In Vitro Transfection Kits. The IRR-MRC5 Transfection Reagent is an advanced proprietary cationic lipids formulation, and it has been developed to provide high transfection efficiency with the IRR-MRC5 cell line. This cell line is a good host for studying mammalian embryonic tissue. When cultured in vitro, IRR-MRC5 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 IRR-MRC5 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

High Density IRR-MRC5 cells Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > IRR-MRC5 Transfection Reagent (Irradiated Fibroblasts) The IRR-MRC-5 cell line originates from lung tissue of a 14-week-old Caucasian male fetus. The primary function of the IRR-MRC-5 cells is to support the growth of other cells. In regards to the nomenclature, the “IRR” label specifies that the cells have been irradiated and will not replicate or divide like other cells. This cell line displays fibroblast morphology and has applications for studying human embryonic lung tissue. High Density IRR-MRC5 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

IRR-MRC5 Transfection Protocol Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > IRR-MRC5 Transfection Reagent (Irradiated Fibroblasts) IRR-MRC5 Transfection Protocol 1. Plate 10,000 - 15,000 IRR-MRC5 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 IRR-MRC5 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 IRR-MRC5 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 > IRR-MRC5 Transfection Reagent (Irradiated Fibroblasts) 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

IRR-MRC5 Transfection Kit Product Details Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > IRR-MRC5 Transfection Reagent (Irradiated Fibroblasts) IRR-MRC5 Transfection Kit Product Details Proprietary 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 > IRR-MRC5 Transfection Reagent (Irradiated Fibroblasts) Data Figure 1. siRNAs targeting Lamin A/C mRNA or non-silencing control siRNA were transfected into IRR-MRC5 cells following the recommended protocol. At 48 hours post-transfection the cells were analyzed by qRT-PCR for Lamin A/C gene expression levels. 18S rRNA levels were used to normalize the Lamin A/C data. Values are normalized to untreated sample. Data are means ± SD (n=4).

Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > IRR-MRC5 Transfection Reagent (Irradiated Fibroblasts) Data Figure 2. Protein expression of Lamin A/C in IRR-MRC5 cells. DNA plasmid expressing Lamin A/C or siRNA targeting Lamin A/C were transfected into IRR-MRC5 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.

IRR-MRC5 Transfection Kit Benefits Leading Developer and Manufacturer of In Vivo and DNA Transfection Kits, Transfection Reagents and Electroporation Delivery Products Products > IRR-MRC5 Transfection Reagent (Irradiated Fibroblasts) IRR-MRC5 Transfection Kit Benefits Pre-optimized transfection protocol for IRR-MRC5 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 IRR-MRC5 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 IRR-MRC5 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