siRNA / RNAi /miRNA transfection Rat IEC-6

Get tips on using FuGENE® 6 Transfection Reagent to perform DNA transfection Mammalian cells - Primary cells Rat pulmonary artery smooth muscle cell (pPASMC)

Get tips on using Oligofectamine™ Transfection Reagent to perform siRNA / RNAi /miRNA transfection Rat - A-10 Cationic lipid based

Get tips on using Xfect™ Transfection Reagent to perform siRNA / RNAi /miRNA transfection Rat - H9c2 Cationic and neutral lipids

miRNA is the inherent gene silencing machinery which can have more than one mRNA target, whereas siRNA can be designed to target a particular mRNA target. By design, both siRNA and miRNA are 20-25 nucleotides in length. The target sequence for siRNAs is usually located within the open reading frame, between 50 and 100 nucleotides downstream of the start codon. There are two ways in which cells can be transfected with desired RNAi: 1. Direct transfection (with calcium phosphate co-precipitation or cationic lipid mediated transfection using lipofectamine or oligofectamine), and 2. Making RNAi lentiviral constructs (followed by transformation and transduction). Lentiviral constructs are time consuming, but provide a more permanent expression of RNAi in the cells, and consistent gene silencing. Direct transfection of oligonucleotides provides temporary genetic suppression. Traditional methods like calcium phosphate co-precipitation have challenges like low efficiency, poor reproducibility and cell toxicity. Whereas, cationic lipid-based transfection reagents are able to overcome these challenges, along with applicability to a large variety of eukaryotic cell lines. When using oligos, the ideal concentration lies between 10-50nM for effective transfection.

Get tips on using FuGENE® 6 Transfection Reagent to perform DNA transfection Mammalian cells - Primary cells Human chondrocytes

Get tips on using Rat IL-6 Quantikine ELISA Kit to perform ELISA Rat - IL-6

Get tips on using Rat IL-6 ELISA Kit (ab100772) to perform ELISA Rat - IL-6

Get tips on using Rat IL-6 PicoKine™ ELISA Kit to perform ELISA Rat - IL-6

ELISA is the most commonly used method of detecting and quantifying the concentration of an antigen in an unknown sample. During the experiment, If you get a weak signal, then make sure reagents are at room temperature before starting the assay. Try increasing incubation times to ensure maximal antibody binding and amplify the signal. Secondly, if you get values above 0 in the negative control indicates a high background signal. Try to consider reducing your antibody concentration and prevent non-specific binding of antibodies by using affinity-purified antibody and suitable blocking buffers. To avoid high well to well variation, do not stack plates during incubation, no bubbles in the plate and wash wells thoroughly to avoid variation.