Since their introduction in the 1950’s immunoassays have been an integral part of research and drug discovery labs. Immunoassay platforms are commonly used to detect and quantify levels of an analyte from a range of complex sample matrices (e.g. human serum, urine, or blood) where an abundance of other proteins and molecules are present. Over the years the immunoassays available to scientists have expanded, leading to the plethora of immunoassay platform options we now have today. Some of the most commonly used immunoassay platforms include radioimmunoassays, western blots, and ELISAs. PerkinElmer offers a range of immunoassay platforms that offer multiple benefits over these more traditional means.
Traditional ELISAs are commonly used in disease research and throughout the drug discovery workflow. While compatible with complex matrices, a colorimetric ELISA is susceptible to the constraint of a narrow linear range for the optical density (OD), which is common to absorbance-based measurements. The smaller dynamic range (~ 2 logs) of the assay decreases the likelihood that your unknown sample concentration will fall within the standard curve and introduces the challenge of testing multiple dilutions from the same, potentially limited, sample. Repeat dilution testing of the same sample also requires additional assay wells and often leads to purchasing additional kits or coating additional assay plates to accurately quantify all your samples. Chemiluminescent ELISAs can be more sensitive, but require vast amounts of antibody, increasing the cost of the assay, and lack signal stability due to the nature of the rapid decline in luminescence over time. Our ELISA-alternative technologies provide not only a wider dynamic range and expanded signal stability, they also offer increased sensitivity and the option for a no-wash assay thus elimination some of the most common limitations of ELISAs.
Similarly, western blotting is an immunoassay platform that is widely used as a semi-quantitative method to determine and compare amounts of protein. These assays can suffer from limited sensitivity and throughput. Because western blotting involves a multistep protocol, variation and errors can occur at any step, reducing the reliability and reproducibility of results. Out western blot alternative immunoassay platforms provide easy-to-use means for protein quantification.
PerkinElmer offers a range of ELISA alternative immunoassay technologies that enable selection of the optimal immunoassay platform for your requirements. From sample compatibility to sample volume, throughput, detectable range, signal window, signal stability, reproducibility, automation compatibility, and desired speed of results, PerkinElmer offers immunoassay technology platforms, pre-configured kits, and customizable reagents for superior results.
Benefits of PerkinElmer Immunoassay Platforms:
|ELISA||DELFIA TRF||HTRF® and LANCE Ultra TR-FRET||Alpha|
|Plate Format||96-wells||96- to 384-wells||96- to 1536-wells||96- to 1536-wells|
|Time to Results||4-24 hours||4-6 hours||1-2 hours||1-2 hours|
|Dynamic Range||2 Log||5 Log||4 Log||5 Log|
|Sensitivity||10 pg/mL||10 pg/mL||10 pg/mL||1 pg/mL|
|Avidity||No||No||Only with streptavidin||Yes|
|Signal Stability||Up to 1 hour||Days - Years||24 hours - Days||Up to 24 hours|
|Multiplexing||No||Yes (up to 3 targets)||No||Yes (up to 3 targets)|
|Reader Requirements||Chromogenic and Chemiluminescence Compatible||TRF Compatible||TRF Compatible||Alpha Compatible|
Flurorophore Cryptate (HTRF)
|Flurophore Embedded Beads|
|Analyze Size||No Limitation||No Limitation||Subject to Forster Radium (9nm)||Subject to oxygen singlet diffusion (200nm)|
DELFIA® TRF is a proven, robust wash-based immunodetection platform with over 20 years of history. DELFIA has a similar assay principle and workflow to that of a traditional ELISA, but with the added benefits of a stable, time-resolved fluorescent signal improved assay dynamic range. DELFIA also offers the ability for multiplexing. DELFIA TRF is compatible with the widest range of sample types, ranging from simple buffers, cell supernatants, and cell lysates to complex biological samples, including tissue, serum, plasma, and blood.
Below is a workflow comparison between a DELFIA TRF assay and a traditional ELISA.
LANCE Ultra TR-FRET
LANCE® Ultra TR-FRET is a no-wash proximity assay technology that combines the benefits of time resolution (TR) with fluorescence resonance energy transfer (FRET) principles by using energy donor and energy acceptor fluorophore labels. It’s everything you love about TRF detection with fastest results and no wash steps. LANCE Ultra TR-FRET is compatible with buffer, cellular supernatant and cell lysate samples.
Below is a workflow comparison between a LANCE Ultra TR-FRET assay and a traditional ELISA.
HTRF® (Homogeneous Time Resolved Technology) is a FRET-based, no-wash technology.
Rapid, homogeneous, easy to use and to automate, allowing extreme miniaturization and easy assay development – over time, HTRF has become the gold reference for time-resolved FRET.
HTRF technology can be used in competitive and noncompetitive formats, performed as cellular or biochemical assays and thus can been applied to a variety of applications including GPCRs, kinases, epigenetics, and quantification of a range of biomarkers including cytokines.
Below is a workflow comparison between an HTRF assay and a traditional ELISA.
Alpha is a bead-based, no-wash luminescent technology. When Alpha Donor and Acceptor beads are brought together through binding to target, a cascade of chemical reactions is set in motion, creating an amplified signal. These versatile beads are conjugated with various biomolecules that enable detection of unique biological events, offering maximum flexibility. Alpha technology is compatible with a wide range of sample types ranging from simple buffers, cell supernatants, and cell lysates to complex tissue, serum, plasma, and other biological samples.
Alpha offers the benefits over ELISAs including:
Below is a workflow comparison between an Alpha assay and a traditional ELISA.