Custom Human IKK-gamma/NEMO/FIP-3 Immuno-PCR Assay Kit, IQELISA®

ELISA Kits|Immuno-PCR Assays

Cell Culture Supernatants
Plasma
Serum

qPCR

96-well Microplate **The PCR plate used is a 0.2mL, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600)

IKK-gamma (
NEM
O/FIP-3) Microplate (Item A)**: 96 well PCR plate coated with anti-Human IKK-gamma (
NEM
O/FIP-3)
Wash Buffer I Concentrate (20x) (Item B): 25 ml of 20x concentrated solution
Standards (Item C): 2 vials of recombinant Human IKK-gamma (
NEM
O/FIP-3)
Assay Diluent A (Item D): 30 ml diluent buffer, 0.09% sodium azide as preservative. For Standard/Sample (serum/plasma) diluent
Assay Diluent B (Item E): 15 ml of 5x concentrated buffer. For Standard/Sample (cell culture medium/urine) diluent
Detection Affinity Reagent for IKK-gamma (
NEM
O/FIP-3) (Item F): 2 vials of a 4x concentrated solution of anti-Human IKK-gamma (
NEM
O/FIP-3) affinity reagent IQELISA Detection Reagent (Item G): 1.4ml of a 10x concentrated stock
Primer Solution (Item I): 1.7 ml vial
PCR Master Mix (Item J): 1.2 ml vial
PCR Preparation buffer (Item K): 1ml vial of 10x concentrated buffer
Final Wash Buffer (Item L): 10ml vial of 10x concentrated buffer**
The PCR plate used is a 0.2mL, non-skirted 96-well plate (ThermoFisher, cat. no.: AB0600)

Prepare all reagents, samples and standards as instructed
Add 25µl standard or sample to each well. Incubate for 2.5 hours at room temperature or overnight at 4°C
Add 25 µl detection affinity reagent to each well. Incubate 1 hour at room temperature
Add 25µL of IQELISA Detection Reagent to each well. Incubate 1 hour
Add 15µL Primer solution 10µL of PCR master mix to each well
Run real-time PCR

The RayBio® Immuno Quantitative Enzyme Linked ImumunoSorbent Assay (IQELISA) is an innovative new assay that combines the specificity and ease of use of an ELISA with the sensitivity of real-time PCR. This results in an assay that is simultaneously familiar and cutting edge and enables the use of lower sample volumes while also providing more sensitivity. The RayBio® Custom Human IKK-gamma (NEMO/FIP-3) IQELISA Kit is a modified ELISA assay with high sensitivity qPCR readout for the quantitative measurement of Human IKK-gamma (NEMO/FIP-3) in serum, plasma, and cell culture supernatants. This assay employs an antibody specific for Human IKK-gamma (NEMO/FIP-3) coated on a 96-well PCR plate. Standards and samples are pipetted into the wells and IKK-gamma (NEMO/FIP-3) present in a sample is bound to the wells by the immobilized antibody. The wells are washed and a detection affinity molecule is added to the plates. After washing away unbound detection affinity molecule, primers and a PCR master mix are added to the wells and data is collected using qPCR. Ct values obtained from the qPCR are then used to calculate the amount of antigen contained in each sample, where lower Ct values indicate a higher concentration of antigen.