*3.4.1. Pipette stations*

reduction of false negative hits, as damaged or dirty pins would usually deliver lower vol‐ umes than anticipated. In dose-response analysis, the quality of the curve fit is highly de‐

A good quality control procedure should provide the transferred volume and the variation associated with the pin set. We implemented a relatively quick and simple procedure using a fluorescent dye (FITC). Prior to the test, the pins are washed as described above. A calibra‐ tion curve is generated of fluorescence intensity as a function of FITC concentration. Using the pintool, FITC in DMSO is transferred from a source plate to several destination plates containing PBS (the use of 4 plates was shown to be sufficient). The average transferred vol‐ ume per pin is calculated using the fluorescence signal of the destination plates and the cali‐ bration curve. Volume variation across the microtiter plate can be readily appreciated by plotting volume against well position (Fig. 5, top charts). The pink and green solid lines rep‐ resent the upper and lower boundaries within 10% CV of the average volume, where outli‐ ers can be clearly identified. The frequency chart (Fig. 5, bottom chart) displays outliers present in 1, 2, 3 or all of the 4 destination plates, and it can be used to identify pins that consistently provide volumes outside a specified range. In the example shown in Fig. 5, pins corresponding to positions A13, B21, D8, F13, K1, N14 and P20 will have to be replaced. De‐ pending on the need, stringency can be adjusted by changing the boundaries as specified by %CV. It is highly recommended to utilize the same freshly prepared fluorescent dye and buffer solutions in all aspects of the protocol. A template for data analysis can be easily cre‐

**Figure 5.** A simple and comprehensive approach to analyze pintool performance. Individual pins can be selected for

pendent on the variability of the data points.

188 Drug Discovery

ated in conventional software such as MS-Excel.

replacement based on consistent variation across multiple transfers.

The automation station is an integral part of any high throughput pipettor, regardless of the type of tips (fixed or disposable) it employs. It typically consists of ANSI/SBS standard com‐ pliant single or multiple deck positions on a stationary or moving platform to hold the lab‐ ware, with a moving arm situated above the platform containing the single- or multichannel pipette head. A major advantage of automated pipettor devices over manual or electronic multichannel hand-held pipettes is the elimination of inconsistency in the transfer process by minimizing human intervention, which also enables high throughput applica‐ tions that are not otherwise feasible. The three major tasks that can be performed with suita‐ ble hardware settings are liquid transfer, cherry-picking and serial-dilution.

For plate-to-plate liquid transfers, 96- or 384-well pipette heads are preferred to work with 96-/384-/1536-well microplates to speed up the process and increase the throughput. While 4-/8-/12-/16-pipette heads can also be used for direct transfer applications, they are primarily used to perform serial-dilutions. On the other hand, a single channel pipette tip is an essen‐ tial component to accomplish cherry-picking tasks.

The speed of an automated pipettor is important for time-sensitive experiments. Especially when performing small volume transfers into microplates, the amount of time spent to transfer liquids in a column-by-column or row-by-row manner may be problematic due to quick evaporation. If the speed of transfer is too slow, some evaporation in the first column or row may be observed before dispensing to the last column or row, causing inconsistent volume across the plate. To avoid evaporation issues during liquid transfers, deck size, pi‐ pettor speed, head type and the transfer volume should be considered.
