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FlowJo Basic and Advanced Tutorial – Save the date!
November 1st 2-5pm – WIMR Seminar Room C2.20.
We will be hosting Jack Panopoulos – FlowJo Application Scientist – as he delivers updates on basic and advanced flow cytometry data analysis using FlowJo & SeqGeq. There have been a number of recently released plugins that there will also be focus on that should interest many.
NanoFCM demonstration – Get your 7-500nm samples ready!
September 19th & - October 15th – WRHFlow Labs - J2.05
Small particle detection/characterisation using flow cytometry has been a hot topic in past years. A number of researchers here are currently focusing on nanoparticles/microparticles/viruses. The WRHFlow team have been interested to see how a ‘novel’ flow cytometer / technology performs in this space of <500nm and therefore have organised a demonstration/trial of the NanoFCM.
We will have company engineers on site who can assist running samples for the purpose of the instrument evaluation on the 19th September 2018. We will then have the instrument for ~1 month for evaluation – with assisted sessions available to researchers to evaluate the instrument. Samples will need to be run by WIMR staff.
We will hold a small presentation from 11am-12pm on the 19th on the L7 meeting room to gain an understanding of the technology that you are welcome to attend.
For access, please see reception if needed.
Sample acquisition is slated for the afternoon - ~12-3pm and we welcome your samples for testing.
Please note as this is an instrument evaluation user data may be used to furnish a study report of instrument applicability. Also any data will need to be provided to NanoFCM for review prior to publication. If you are interested in the instrument I would suggest evaluating it with a sample of interest, especially while the engineers are here on site. Further instrument information can be obtained here - https://www.nanofcm.com/en/Product/NanoFCM/117.html
BD Symphony – 8 laser, high sensitivity flow cytometer now installed, training for high parameter panel users.
August 22nd - 11am & 2pm - J2.07
September 4th – 3pm – J2.07
September 21st - 3pm - J2.07
Please RSVP with your intention to attend to wrhflow@sydney.edu.au
View configuration_V1 for panel design here
View configuration_V2 for panel design here
CantoII HTS installed – Time to think about preparing samples in 96 well plates?
All our flow cytometers have now been upgraded with high throughput capabilities. That’s a total of 5 high throughput sampling robots in the flow cytometry facility.
Benefits of sampling from a 96 well V bottom plate include minimised loss of cells between washes, 8-12 samples at a time using a multichannel pipette, better washes as virtually all buffers are aspirated in a 96 well V bottom, and robot acquisition!
If you require training for using the HTS please submit a training request through PPMS.
If you would like a 96 well V bottom plate, these can be ordered through PPMS for $6 each.
BD Symphony install - First Symphony A5.2 (not A5.1) install worldwide!
You may notice a number of engineers in the flow labs this week with the BD Symphony installation/testing underway. Geoff Osborne (BD SORP director) and the team will be busy installing the world’s first updated A5.2 which has a revised detector configuration and new electronics.
We have scheduled in two initial training sessions on the 22nd of August 2018 @ 11am & 2pm (with others to be announced). If you would like to attend this initial training please RSVP to wrhflow@sydney.edu.au for these sessions (we will limit them to 5 each session).
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Failed laser teardown!
Curious what happens inside a diode laser like the below used on the Fortessa? We were un/lucky enough to have had the red laser fail on the Fortessa in August 2018 and so we did a laser teardown.
Lasers come in many shapes/sizes/forms. A very popular choice these days are diode lasers. They are cheap, reliable and come in a range of laser power outputs. The red laser on the Fortessa was one of these.
Solid state laser - Coherent CUBE 639nm 40mW laser
These red lasers have a 'cleanup' bandpass filter to ensure only ~639nm light exits (the little circle lens), and a physical shutter operated by the lever (right)
Simple to open (but tricky, don't forget the hidden screws under the CUBE sticker). The unit pulls apart easily. The main control circuitry, diode, and heatsink are positioned on a nicely CNC'ed solid block of steel (middle).
A close up of the motherboard and diode.
The various boards pull apart easily. We can see the communications board - with the USB connection. The board with the golden connector is for modulating the output. The other circuitry needed for a stable laser include a temperature / emitted light / current draw sensor and control logic. Next to the diode laser is the thermal controller for the heatsink (it can heat and cool the diode). The beam shaping prisms can be seen mounted on the heat sink. This model produces an elliptical beam.
The laser had way more than the average life expectancy for these lasers. This laser had >10000hrs and the diode had failed.
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2018 Tips and tricks competition winners announced!
Congratulations to Kirstie Bertram and Nicole Fewings for winning the WRHFlow Tips & Tricks competition.
Kirstie’s entry highlight a number of important points to consider including antibody clone / manufacturer differences when selecting antibodies for a panel, while Nicole’s entry demonstrates the benefits of titrating not only antibodies but viability dyes. See the/wiki/spaces/DMT/pages/400916724for winning entries.
Instrument bookings...
In the last 6 months the 4 flow cytometry analysers located at WRHFlow have seen over 100 different users resulting in over 2000 hours of actual collective usage, with about 1/4 of usage being after business hours. These numbers are wonderful and depict an increasing usage of the instruments.
There are 2 statistics however that are of considerable interest to me.
41% of the time booked on instruments is not used.
50% of the time used is not booked.
Please users, remember to cancel sessions that can not be used as soon as possible, as this will undoubtedly assist other users who may be staying later to run samples. This will also allow WRHFlow to remain not charging for unused booked time. The latter which occurs at other facilities.
Fortessa HTS upgrade!
The Fortessa now has the hardware required for acquisition from a 96/384 well plate. We have organised an en-masse training session on the 12th of July @ 9am for any one who would like to use it.
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Notice - SIP sheath for the LSRII and Fortessa
The LSRII and Fortessa must have SIP sheath covers installed to prevent dripping of samples.
- The SIP sheath cover is installed by default on both instruments.
- The SIP sheath cover can be removed (but must be reinstalled immediately for a different sample or following your session) for the following samples only if -
- Bead samples are immediately to be acquired, OR,
- The samples have been inactivated by fixation. An example protocol for sample fixation could consist of 20 minutes exposure @ 4°C to a flow cytometry specific commercial fixative containing paraformaldehyde (example buffer set & protocol). Other protocols that result in fixation of samples are also valid, OR
- The samples are being acquired from a HTS device.
- A properly installed SIP sheath cover will look slightly thicker than without the SIP sheath (compare with below photo) and should not drip liquid when the instrument is on run. To remove the SIP sheath on the LSRII or the Fortessa follow these instructions. To replace it, the order is reversed.
- To reinstall the SIP sheath cover follow the reverse instructions. The above applies to the LSRII & Symphony.
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WRHFlow have instigated some automatic data syncing scripts for users who would like to try them out. Unless your data should not be synced to the WIMR server, when exporting your data, export it as FCS3.0 format, with linear format selected to 'D:\BDExport\FCS' (as this folder will from now be automatically synced to the WIMR server). If you would like to copy your data elsewhere you can also copy it from this location. Your data should be able to be accessed from the Scientific Platforms networked drive within WIMR, and via a username/password for non-WIMR users via Dropbox/Onedrive if desired (email wrhflow@sydney.edu.au to setup an account).
Drop off your sample for multiplex analyte analysis?
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