How to use the Mosquito
Because of the speed of operation, there is no booking system on the Mosquito. However, the screens (14 different screens pipetted into 96-well plates) are stored in a locked fridge. To request plates, first log their use on Sharepoint, then send an email to James E. Mansfield and Marc Morgan who will take the requested plates out for you within 24 hours and put them in the 4°C incubator. This done, come up to Flowers 5.20 with a suitable amount of protein (18 µl for one set of 100 nl drops) and you're ready to go.
Before the experiment
- Turn on the laptop next to the Mosquito and log on to user account (no password), turn on the Mosquito (switch behind on the left) and start the Mosquito software.
- Click Yes when asked to initialize the Mosquito.
- Click the Open File icon and select the appropriate program.
- Understand the program and verify that it will do what you want it to do.
A large number of programs have been written for the Mosquito, most of them for 96-well MRC plates (called MD2 internally). They are organized by theme:
- One_Drop – For one-drop experiments, protein goes into the thin strip in the middle of the stage.
- Two_Proteins – For experiments with two different proteins or one protein with two different ligands, protein goes into the two leftmost strips on the four-strip holder.
- Two_Ratios – For experiments with one protein but two different drop ratios, protein goes into the strip in the middle of the stage. Otherwise, these protocols are identical to the Two_Proteins protocols.
- Two_Ratios_fast – To set up faster one-protein-two-drop-ratio experiments, the crystallization solution is added to both drops with one needle. Tips are changed "Between Transfers" in the Aliquot steps and not at all (Multidispense) in the Copy steps. Protein goes into the strip in the middle of the stage.
- Additives – To do additive screening with the Mosquito, additives should be provided in 96-well round-bottom plates, which can be booked on Sharepoint.
- LCP – Cubic lipid phase experiments can be set up with SwissCi LCP plates.
- Microseed_Screening – The Multiaspirate feature is used to aspirate seed stock and crystallization solution for combined dispensing.
- MRC_Maxi – Setting up MRC Maxi plates (48 wells) is not recommended because the required volumes often exceed the maximal capacity of the Mosquito tips (1.2 µl).
- Low_Profile – We have protocols for CrystalX, Mitegen In-Situ-1 and Greiner XLT plates.
- Unusual_Protocols – This is the place for under-oil crystallization and other weird stuff.
- User_Defined – Users should save their modified protocols here
Running the experiment
- Place a sample strip into the holder you want to use (middle holder for one-protein experiments, else left holder) and cover with magnetic cover.
- Add protein to each well, as required by the chosen program. Avoid bubbles.
- Carefully remove the tape from a prepared screen and place screen in rightmost position on the sample stage, the label facing you.
- Click Run to start the program.
- Make sure the used needles are collected into the yellow bin. Avoid touching the needles with your bare fingers. They are sharp and might be contaminated. (For example, cacodylate buffer contains arsenic.) Gloves are in the drawer closest to the exit.
- When protocol is finished, remove plate and tape it shut. Use the squeegie to ensure a tight seal.
- Store plate in one of the big incubators (4°C or 20°C).
- Repeat until you're done with all your plates
After the experiment
- Remove and discard used sample strips.
- Park the sample stage in the middle.
- Sleep computer (by pressing Off button) and close lid.
- Turn off Mosquito.
Please leave the facility in a tidy state and report any errors to the facility manager.