Frequently Asked Questions

 

General
What are the major differences between the DropArray Microplate and a conventional microplate?

Rather than using physical wells, DropArray Microplates are completely flat including a series of 96 or 384 hydrophilic spots. A hydrophobic surface occupies the entire flat surface area of the plate except for the spots. These spots form an array that conforms to the well layout of the ANSI/SBS 4-2004 standard. Using standard microplate pipetting devices, liquid dispensed onto these spots form droplets that remain in a fixed position, even when the plate is shaken or inverted. Plastic walls surround the entire flat surface, which serves to contain washing buffers that are added in bulk to the plate. Unlike a standard microplate that requires dispensing and aspiration from each well, bulk liquids like washing buffers are added to or removed from the entire DropArray Microplate at once. By reducing turbulence in wash steps, far fewer cells and beads are lost from the assay droplets, even with suspension cells and non-magnetic beads. Evaporation is controlled using an Evaporation Control Lid which provides a humidified environment and minimizes evaporation of the droplets.

What are the distinct advantages of DropArray Microplates over conventional microplates?

DropArray Microplates enable simple and easy miniaturization of assays at 2-4 microliters for the 384-well format and 5-20 microliters for the 96-well format because of their unique wall-less design. The wall-less feature eliminates capillary action, the biggest barrier to miniaturization with conventional microplates. Furthermore, the wall-less feature allows extremely gentle washing by liquid exchange, which is different from the harsh, convection-based washing performed by a microplate washer with an array of nozzles.

Luminex® Assays
What are the benefits of using DropArray DA-Bead plates for running Luminex assays such as Milliplex®, Bio-Plex® and Procartaplex® assays?

A DropArray DA-Bead plate reduces the consumption of both samples and beads to 5 microliters each, saving 80% in reagent costs while using the same 96-well format. Including the cost of a DropArray DA-Bead plate, overall assay costs are generally reduced by 70%. Furthermore, the consumption of only 5 microliters of sample per plate droplet makes DA-Bead plates ideal for precious samples such as mouse sera and human tears.

Wouldn't dispensing of 5 microliters of sample and bead reagent increase my CV compared to dispensing 25 microliters on a standard microplate?

Running Luminex assays at 5 microliters on DropArray DA-Bead plates does NOT increase CVs, in spite of the fact that higher CVs are inevitable when dispensing small volumes like 5 microliters. It is known that the higher CVs for Luminex assays stem from a combination of steps, mostly from dispensing, washing, and Ab-Ag binding on the bead surface. While the DropArray system may produce higher CVs purely due to dispensing in smaller volumes, it achieves lower CVs in the washing and Ab-Ag binding steps due to uniform and gentle washing and incubation of the plate WITHOUT shaking. Therefore, users who run Luminex assays on a DropArray DA-Beat plate will observe CVs at least comparable to, if not better than those obtained with conventional microplates.

Does a DropArray DA-Bead plate change my current Luminex assay workflow on a conventional microplate?

No. The entire workflow of a Luminex assay on a DropArray DA-Bead plate is identical to that of a conventional microplate except for two steps. The first is the incubation of the plate. A DropArray DA-Bead plate does NOT need shaking. It is incubated in a static position which is also more convenient. The second is the transfer of beads from a DropArray DA-Bead plate to a conventional V-bottom plate such as a PCR plate. This extra step takes 5-10 min. You can watch a video of the entire assay run on a DropArray DA-Bead plate at http://www.curiox.com/videos.html. Curiox's team is currently optimizing the direct reading of a DropArray DA-Bead plate using a Luminex reader.

Is it easy to use a DropArray DA-Bead plate, particularly in dispensing? Doesn't it require special skills as it has no walls?

A DropArray DA-Bead plate is as easy to use as a conventional microplate because the hydrophilic spots on the plate attract reagents while the hydrophobic surface between the spots repels reagents. A dispensed droplet adheres to a spot and does not come off during the typical handling expected for a plate. When dispensing reagents at small volumes, a DropArray DA-Bead plate is much easier to use than a conventional microplate because, for example, even 2 microliters of reagent is distinctively visible as a droplet on a plate.

Why doesn't miniaturization of a Luminex assay on DA-Bead plates affect the sensitivity? Wouldn't the reduction of sample volume decrease the sensitivity as it has a smaller amount of a target analyte?

The miniaturization of a Luminex assay does not affect the sensitivity, and this has been proven by numerous users of the DropArray system. In a Luminex assay, the ratio of sample volume to the number of beads is determined at 25 microliters of sample for ~2,000-2,500 beads. When an assay is miniaturized by 5-fold on a DA-Bead plate, both the sample and the number of beads are reduced correspondingly. It leaves the same amount of a target analyte available per bead, leading to the same sensitivity in general.

Are there enough beads when a Luminex assay is reduced to 5 microliter bead solution on a DA-Bead plate?

Yes. In a 25 microliter bead solution, there are ~2,000-2,500 beads per analyte. A 5 microliter bead solution contains ~400-500 beads per analyte, which is still excessive for the number of beads required for reading.

Does the whole-plate washing of a DropArray DA-Bead plate cause cross-contamination between droplets?

No. Numerous users have proven that there is no detectable cross-contamination between droplets. There are two possible pathways of cross-contamination on a DropArray DA-Bead plate. One is the transfer of beads between droplets during washing. It has been proven experimentally that beads do not move out of a droplet during washing. For example, one bead placed in a droplet remains in the same droplet even after 10 washing cycles. Such excellent retention of beads during washing is due to the gentle washing by liquid exchange which is unique to the DropArray system.

The other possible pathway of cross-contamination is the transfer of a sample between droplets during washing. This has also been extensively tested and validated to confirm that there is no cross-contamination. The washing performed by the DropArray system is based on substantial dilution of samples and reagents on a plate by introducing a bulk amount of wash buffer repeatedly during a short period of time. For example, upon the completion of a 3x washing process, there's zero chance of a sample moving to a neighboring droplet and residing in that droplet in a way that affects the outcome of an assay.

How much wash buffer and washing time are needed to wash plates on the DropArray Washing Station?

A DropArray Washing Station typically needs ~110 ml for a 1x washing cycle. A 1x wash cycle and 3x wash cycles need ~2 min and ~4 min respectively, including 30 seconds of built-in rest time needed before starting the next wash process for precipitating beads onto the surface of a plate.

Are DropArray Washing Stations automation-friendly?

Yes. The DropArray HT Washing Station (HT200) has a PC-HT200 RS232C port for communication with automation controllers.

Cellular Assays
Do we need any specialized dispensers or imaging systems?

No. The DropArray platform allows seamless use on any conventional dispenser, gripper, stacker, reader or imager.

What are the cell growth parameters on a 384-well DropArray DA-Cell plate?

The DropArray DA-Cell plate can be used with a low confluence of 80 cells per mm2 to a very high confluence of 630 cells per mm2. For first time users, we recommend 250 cells per mm2. The cell growth rate on DA-Cell plates is no different compared to a conventional microplate.

How long can the cells survive on DropArray DA-Cell plates? How do I perform media exchange?

Depending on cell-line and assay requirements, cells can survive up to two weeks without media exchange. Media can be aspirated manually or washed off using DropArray HT/LT Washing Stations before applying fresh media.

Do the DA-Cell plates come coated?

DropArray DA-Cell plates are tissue culture-compatible. Like a conventional tissue culture plate, DA-Cell plates can be easily coated with DNA, poly-lysine, Collagen, fibronectin, silane-compatible substrates, or any other cell culture coatings. In addition, pre-coated plates for different cellular assays are available.

Milliplex, Bio-Plex, Procartaplex and Luminex are either trademarks or registered trademarks of eBioscience and Luminex Corporation respectively in the US and/or other countries. Curiox DA-Bead plates are not endorsed by the Luminex Corporation.