Kinetic Studies

kinetic assays using plate read and stacker

There is a need for kinetic assays to be run on low-cost automated equipment. PAA have teamed up with Molecular Devices to produce a fully automated system for LaTrobe University, Melbourne, Australia.

The Problem

SYTOX Green stain provides a quantitative single-step dead-cell indicator. Using this, bacterial viability can be monitored over a long time frame.  This is usually performed as a kinetic assay, in microtitre plates, with a fluorescent plate reader allowing a large number of experiments to be conducted. The problem is running the experiment over a time period of 24 hours. If only plate is being used, it can be left in the reader. However if multiple plates are to be run, the task becomes more difficult requiring the plates to be cycled through the reader at prescribed times.

PAA solution

  • Use a Molecular Devices SpectraMax plate reader and StakMax stacker
  • Use Overlord from PAA to control the work flow
  • Use Microsoft Excel and a program written by PAA to collate and display the data
What can we do now, we couldn’t do before?

SYTOX kinetic studies are time consuming and difficult to perform because of the time delays between plate reads, keeping track of the plates to read in the plate reader and collating the data.

PAA in association with La Trobe University (Australia) and Molecular Devices have developed a low cost and reliable solution to this technique.

System

The hardware for this system consists of:

  • Molecular Devices SpectraMax plate reader
  • Molecular Devices StakMax plate stacker (with optional barcode reader for sample tracking)

The software consists of:

  • Overlord laboratory automation control software
  • Microsoft Excel
Process
Prepare the set of plates required for the kinetic assay, the Molecular Devices StakMax can handle 20 plates.

The requirement is to run this series of microplates in the plate reader once a day (or any other time delay). Overlord controls the following sequence of events

  1. determine the read time interval
  2. get a plate from the input stack, read the barcode
  3. open the plate reader plate drawer, load plate
  4. close the plate reader drawer
  5. run the required plate reader method
  6. when finished open the plate drawer
  7. get the plate from the plate reader
  8. return the plate to the output stack
  9. repeat steps 1-8 for all plates
  10. re-stack the plates from output stack to input stack
  11. wait the required time interval
  12. run steps 1-11
  13. run steps 1-12 until all the data has been collected
Overlord3 control software
overlord-mockup-cropped
Overlord handles:
  • Control of the instruments
  • Running the process
  • Timing the delay between read n and read n+1
  • Collating the barcode data
  • Passing the data to Microsoft Excel for data management and display

more information: Overlord

Handling results

The system records the data as the run proceeds and at the end of the run, the data is available for review. One data file is produced for each plate in the run.

The system allows replicates to be placed on the plate. A plate map in Excel allows the system to know where each sample is on the plate.

These results are then averaged to give a result for each sample on the plate. Each sample is summarised on a separate tab of the results workbook.