Preparing the samples

 

To minimize evaporation, prepare the samples immediately before run time or keep the plates lidded until you run the protocol.

 

When preparing the samples, you must:

•	Remove macromolecular particulates before the samples are loaded onto AssayMAP cartridges.

•	Adjust the buffer composition to optimize the binding conditions.

•	Determine the volume of samples to load on the AssayMAP cartridges.

•	Transfer the samples to the microplate you want to use for the protocol run.

Removing macromolecular particulates

Make sure the samples are free of macromolecular particulates, such as large protein aggregates and cellular debris. If the samples have passed through a chromatography column, they should be free of particles and can be loaded on to an AssayMAP cartridge. Samples that might contain particles should be filtered ideally through a 0.45‑µm filter or centrifuged at a high g-force before loading on an AssayMAP cartridge.

 

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Adjusting the buffer composition

Depending on the type of fractionation you are performing the sample should be in a buffer compatible with the stationary phase. For peptide fractionation using SCX cartridges, optimum performance is typically achieved under low salt and low pH conditions. For low-pH peptide fractionation using C18 and RP-S cartridges, samples typically should have a pH of < 3. For high-pH peptide fractionation, optimum performance is generally achieved when the pH > 10.

 

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Determining the volume of sample to load

AssayMAP cartridges can accommodate volumes up to 250 µL.

 

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Transferring the samples to labware

Setting up the sample microplates

Before transferring the samples, you should plan the layout of the samples in the microplate. Consider the following:

•	You can process 1 to 96 samples in parallel. The AssayMAP Probe Syringes always move in unison. The position of samples in their microplate dictates the positions needed for cartridges in the 96AM Cartridge & Tip Seating Station, and the reservoirs for solutions at the other deck locations.

•

When you use only a subset of wells in a sample microplate, fill the same subset of positions at all the deck locations. The default settings for the AssayMAP platform assume that samples will be arranged in multiples of 8 in a column-based configuration. The Bravo Platform applies differential pressure to seat cartridges based upon the number of full columns of cartridges. Therefore, to achieve proper cartridge seating entire columns must be used to guarantee optimum performance.

•	If the number of samples you have is not a multiple of 8, use the AssayMAP Resin-Free cartridges to fill the empty column and row positions. This will prevent liquids from dripping on the deck or being dispensed on the deck during the cup washing step.

Figure. Sample microplate layout and reservoir recommendations

 

Transferring the samples to the microplate

You can transfer the samples to the microplate that is supplied with the AssayMAP kits. See Starter kit, cartridges, and labware.

 

A small reagent volume excess is required in all labware types to ensure proper volume transfer.

 

An excess (overage) volume ensures that a microplate well or column does not fully deplete, which would result in aspiration of air into the syringes and then into the cartridges, compromising performance. For the specific pipetting overage requirements of a given labware type, see the Labware Reference Guide in the Literature Library page. Modify this excess volume accordingly if evaporation of volatile solvents is a concern.

To transfer the samples to the microplate:

1	Run the provided Reagent Transfer utility to transfer the samples. For instructions, see Reagent Transfer v2.1.

2	If necessary, centrifuge the sample labware to remove bubbles.

 

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