An original design by Yoon Lee, Greenhouse Lab at the University of California, San Francisco, Division of Infectious Diseases.

Version 1.3

Problems/complaints this device addresses:

Current microarray assay systems require the user to process each well in a one-by-one fashion. It is done this way for various reasons. The problems arising from processing each well as a single-channel include the following:

1. Sample variability. Duration of exposure to the reagents, namely 2° antibodies and fluorophores/probes, will differ in room temperature during aspiration and washes. A great level of care is required when processing microarray slides in order to prevent scratching of the binding membrane but also to get the aspirating tip close enough to fully aspirate as much of the well’s volume as possible. However, there is a trade-off between this care and the time difference of exposure between the first and last slides to be processes. Additionally, one must also consider that pads are drying out in the moments immediately following aspiration, which can cause significant increase in background noise when imaging.

2. Low throughput. As mentioned in #1, risk of damaging the membrane is significantly higher when using a multi-channel pipettes, and slides bound with antigens are very expensive. Therefore, users resort to using single channel pipettes.

3. As a result of issues related to #1 and #2, users are typically limited to processing only up to four slides at a time. This is due to the amount of worked required for each slide in protein microarray assays. 

4. Also as a result of #1 and in concert with #2, the inability to quickly remove the thin layer of samples of reagent (i.e. human sera, secondary antibodies, E. coli lysate, blocking buffer, etc.) left on membranes after aspiration requires numerous sessions of incubation with wash buffer on a shaker throughout the assay. Not completely removing that thin layer will leave assay components to bind to membrane targets.

Consider that each of these issues are greatly amplified when processing multiple slides simultaneously.

Solutions/benefits to each of the above in order:

1.  The device is aspirated via a single channel connected to sixteen separate, isolated channels. Only a single tip of the aspiration equipment of choice is required. The distance between the exit point and the entry point for each channel are identical despite each well’s different location; This also means that the volume of each channel is also identical, and each well is subjected to about  8.55x10^2 mPa vacuum force using a standard molecular biology lab bench vacuum.

2.  Naturally, aspiration only once per slide will save time. The time that is saved becomes increasingly meaningful when processing multiple slides in tandem. 

3.  Again, time saved. But also keep in mind that this assay, which usually involves two days, might have to be repeated hundreds of times. LazyRA has the potential to keep users happy and motivated in the lab as it can be a laborious and attention intensive process; Mistakes are rarely reversible in protein array assays.

4.  The narrow vacuum nozzle slit at the bottom of each well, leading to individual channels, span 6.5mm in width and just 100 microns in height. It is placed at the very bottom of each well, juxtaposing one side of a membrane. This allows effective removal of stubborn thin layers of fluid every time one aspirates. 

Overall, using LazyRA will reduce variability in treatment of each sample. We have yet to try using our liquid handling robot to automate the processing of slides.  Theoretically, using the OT-OneS, one could process 24 protein microarray slides, resulting in a six-fold theoretical increase of throughput in addition to reduction in treatment variability.