96 well bio dot

Principally, the LFIA performance was demonstrated by adding a series of diluted (1 × PBS) purified virus proteins (SCSMV) [10 µg/ml to 10 pg/ml]. The intensity of the test line was then image captured to establish a standard calibration curve. For that, 30 µl of the purified virus protein sample was dispensed on a sample pad, and allowed to react with AuNP-immunoconjugates for one minute (pre-incubation), then 50 µl of 1 × PBS was added to the sample pad to flow the conjugate contents towards the wicking pad. Pre-incubation might help to improve the detection sensitivity as well. Generally, the visibility of the test line and control line is defined as a positive, and the absence of a visible test line define as negative. With the purified protein sample, the absence of signal was recorded as ultra-low signal. “The presence or absence of a control line respective to the assay is correct or false”. Similarly, the duplex assay was performed with the standard virus protein dilutions, and resulting images were captured for further analysis. Relative sensitivity of this LFIA was also determined through the standard Dot blot assay with SCSMV (1:500) antiserum (96 well Bio-Dot, Biorad) with alkaline phosphatase enzyme conjugates (1:2000) and NBT/BCIP substrates⁴⁴ . The output of colour intensity was image captured and measured to correlate with the enhanced nano-LFIA results.