Thursday, January 30, 2014

ELISA with 100 fold improvement in detection limit & 100 fold less sample volume

My first paper on microfluidic work has been published in Analitica Chimica Acta. This was the first project I worked on for my PhD work (actually the very first one didn't work. That was my advisor's crazy idea he wanted me to give a try.  We abandoned that later on). 

In this article, we have demonstrated a novel approach to enhancing the sensitivity of enzyme-linked immunosorbent assays (ELISA) through pre-concentration of the enzyme reaction product (resorufin/4- methylumbelliferone) in free solution. 

Highly sensitive analytical techniques are required to estimate small amounts of disease markers (e.g., antibodies/antigens) in bodily fluids in order to detect the onset of dangerous diseases like cancers at their early stages. Here is the abstract of this paper. 

The reported pre-concentration was accomplished by transporting the resorufin/4-methylumbelliferone molecules produced in the ELISA process towards a high ionic- strength buffer stream in a microfluidic channel while applying a voltage drop across this merging region. A sharp change in the electric field around the junction of the two liquid streams was observed to abruptly slow down the negatively charged resorufin/4-methylumbelliferone species leading to the reported pre- concentration effect based on the field amplified stacking (FAS) technique. It has been shown that the resulting enhancement in the detectability of the enzyme reaction product significantly improves the signal-to-noise ratio in the system thereby reducing the smallest detectable analyte concentration in the ELISA method. Applying the above-described approach, we were able to detect mouse anti-BSA and human TNF-a at concentrations nearly 60-fold smaller than that possible on commercial microwell plates. For the human TNF-a sample, this improvement in assay sensitivity corresponded to a limit of detec- tion (LOD) of 0.102 pg mL1 using the FAS based microfluidic ELISA method as compared to 7.03 pg mL1 obtained with the traditional microwell plate based approach. Moreover, because our ELISAs were per- formed in micrometer sized channels, they required sample volumes about two orders of magnitude smaller than that consumed in the latter case (1 uL versus 100 uL). 

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