Traditional hand transfer methods of bulk reagents into microtiter plates are slow and prone to human error. This technology more than doubles the efficiency of the process and helps eliminate that error. This technique can help users understand operations performed in microplates, reduce opportunity for errors, and increase user efficiency during manual pipetting operations.
This technique will positively impact outcomes in pre-clinical and clinical settings by removing and reducing mistakes that can be costly and even life-threatening if not corrected. This open source system can be easily adapted for other applications, such as microplate copy inspections, to eliminate inconsistencies in either compound or bioassay placement, for chemistry and biological needs. Demonstrating the sample transfer procedure will be Lina Deluca, a Compound Manager from the lead identification laboratory.
To set up a semi-automated plate-to-plate sample transfer, use a spreadsheet editing application to generate a CSV file containing source and destination plates, labeling the columns source barcode, destination barcode, source well, destination well, and transfer volume. Under the columns, include one row in the file for each desired pipetting operation indicating the alpha-numeric barcode of the source mircoplate, if there is one, the alpha-numeric barcode of the destination microplate, if there is one, the alpha-numeric row and column identifier for the well to be pipetted from in the source plate, the alpha-numeric row and column identifier for the well to be pipetted from the destination plate, and the volume to be transferred from the source well in the source barcode column, to the destination well in the destination barcode. Open the Light Guide Program to open the microplate assistive pipetting light emitter plate-to-plate graphic user interface application and click Select cherrypick file.
In the file browser window, open the CSV file. The application will parse the first row of the CSV file and illuminate the corresponding wells in the source and destination plates. Use the Previous well"and Next well"buttons to traverse the CSV file as desired.
The graphic user interface will highlight in gray any rows that have been previously illuminated and highlight, in brown, the currently active rows. Perform the pipetting operations as needed to transfer samples between the source well of the source plate, to the destination well of the destination plate. In addition to the user interface, the plate barcodes can be verified via the LCD displays attached to the illumination panels.
Then click Next well"to continue until the end of the CSV file is reached. To load a new CSV file, click Select cherrypick file"at any time. To exit the program, click the red X"at the top right corner of the interface.
For multi-well illuminations, open the microplate assistive pipetting light emitter serial delution application and specify the desired titration mode, plate density, and start rows or columns. Use the Next"and Previous"buttons to navigate through the rows or columns in sequence from the initial start row or column, to the last row or column in the plate. Each time the Next"or Previous"button is clicked, the light panel will illuminate the corresponding LEDs over the microplate.
Continue until the end of the titration sequence is reached before exiting the program. To visually simulate an assay, place a microplate into the portable light guide. The light guide contains a battery and all of the electronics necessary to be used independently of a computer to allow the guide to be used in a handheld mode that can be controlled with built-in push buttons to toggle between demonstration modes.
Use the power toggle switch on the portable light guide enclosure to power the system on and select the appropriate portable light guide mode for the experiment. In the high-throughput screening demo mode, use the right push-button switch at the top of the portable light guide to toggle through the sample illumination patterns. The wells illuminated with red color simulate the reagent dispense of an assay, for example, cells suspended in media.
The wells illuminated with the yellow color simulate detection reagent addition. Next, an excitation light source is simulated with all wells turning blue, followed by simulation of emitted light. The first and last column of wells are illuminated in green, and the middle sample field columns are illuminated in blue to indicate the plate being read on a microplate reader.
Random wells in the sample field will also have green color of varying intensity to represent hits. To toggle the light guide between the high throughput screening demo mode and the titration demo mode, push the left push-button switch. When the light guide enters the titration demo mode, all of the wells in columns three and 13 will be illuminated with the yellow color.
Pressing the right-most push-button switch will illuminate the subsequent columns in sequence. When the push-button is pressed after columns 12 and 22 are reached, the wells in columns four through 12 and 13 through 22 will be illuminated in a decreasing intensity of yellow to represent the titration. For artifact illumination, place the microplate into the portable light guide and press the left-most push-button switch two times to switch the light guide to illumination mode.
Use the right push-button to toggle between a set of predefined colors as needed for the application. The light panel will turn all of the LEDs on in red, blue, green, orange, white, violet, yellow, and indigo in sequence with each push of the right button. Optionally, a camera or smart phone can be used to photograph the illuminated plate for record-keeping or documentation of the work.
The transfer of samples between plates and the preparation of a serial dilution can be accomplished without the concern of distraction or losing track of what pipetting operations remain. The MAPLE platform can be used to help illuminate the microplate to identify potential artifacts, such as precipitate, empty wells, partially filled wells, or air bubbles. Users can then take measures to ameliorate samples before providing them to downstream laboratory processes.
In this representative experiment, the work list consisted of 49 pipetting operations from two 384-well source microplates containing a random assortment of colored dyes that spell Jove in a single 384-well destination microplate. The layout of the wells in the destination plate confirm that the user pipetted into the correct wells. The color pattern of the wells in the destination plate can be used to identify errors for which the user did not pipet from the correct well of the source plates.
Results from this head-to-head test show an average time-saving of 50%when users performed this test using MAPLE versus an offline printed work list. Further, the error rate of the plates created using MAPLE was 0%for all of the users, while a 6%error rate was observed for one novice user using a work list for the sample preparation task. Drug synergy formulations require a 2D matrix preparation based on a serial dilution by column for the first drug and a serial dilution by row for the second drug.
We are investigating the use of LED light panels to illuminate microdroplet dispensing. By strobing LED light, we can capture images of liquid samples for quality control and analysis. This instrument can reduce a user's exposure to hazardous materials and agents by limiting their interaction time and facilitating appropriate reagent usage.
