Proximal Correlation Fluorescence Sensing for HTS Microarray Imaging

The motivation behind this project is that normally to run microarray you need work with 2 different types of instruments. The first being an being an instrument that is used for microarray fabrication. The second being an scanner that generates a high resolution image of the slide. These are normally classified as being capital equipment purchases (ie., items greater then $20K per unit). Added to the cost of array substrate surfaces, as explained in the application note describing how to develop a recycleable bioassay surface, it is clear that despite the potential of conserving high cost biological material and for increasing the throughput using the microarray format, the materials are still too high for many research labs.

So this was project was developed to see if is possible to combine the scanning process with the fabrication process thereby using one instrument to do both tasks. In this example, demonstrated is a proximal correlation fluorescence sensing technique that involves measuring fluorescence microarray spots that are positioned at a sufficiently defined distance from the laser beam diameter. This allows for the spots having fluorescence dye to be consistently excited from the dissipated light from the laser beam diameter. Using this trick, there then generating one high resolution image that involves doing computational intensive image processing and filling data storage, this involves generating many very small pictures that cropped for image processing process where the spots are located as the gantry head containing microscope and light source moves.

It is a very straightforward imaging technique that is fast, quantitative, accurate and scaleable for high throughput. It is also portable and can be configured on standard microarray fabrication instrumentation and gantries like repstraps. This data compares how you can set up a laser and low light sensitive camera to a standard photomultiplier tube confocal microarray scanner.

Video uScope

PMT Laser Scanner

Comparative Data - % Signal Saturation Titration Study

Dye Conc Video uScope
100mW 532nm
PMT Scanner
%80L %55PMT
PMT Scanner
%80L %65PMT
PMT Scanner
%80L %75PMT
3.3uM Sat: % 91.338
CV: % 1.113
Sat: % 96.480
CV: % 2.519
Sat: % 97.083
CV: % 0.875
Sat: % 96.410
CV: % 0.907
820nM Sat: % 91.999
CV: % 2.712
Sat: % 78.905
CV: % 12.237
Sat: % 92.263
CV: % 5.910
Sat: % 97.274
CV: % 1.063
200nM Sat: % 70.504
CV: % 10.892
Sat: % 25.021
CV: % 24.004
Sat: % 69.649
CV: % 17.767
Sat: % 85.148
CV: % 4.104
50nM Sat: % 33.240
CV: % 19.797
Sat: % 5.517
CV: % 24.906
Sat: % 16.839
CV: % 25.133
Sat: % 43.450
CV: % 24.606
13nM Sat: % 18.136
CV: % 19.958
Sat: % 1.639
CV: % 28.856
Sat: % 5.426
CV: % 27.995
Sat: % 13.986
CV: % 28.827

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