American Family Children's Hospital
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Equipment

Introduction

Training

Analysis Computers

Benchtop Analyzers

Sorters

Microscopes

 

Introduction

Please review Flow Cytometry - Using the Facility before making an appointment. Use iLab to schedule appointments for all Flow Lab equipment.

 

You MUST attend the Introduction to Flow Cytometry lecture series before signing up for any training on any cytometer. (Training on microscope is excluded.)

 

New as of Monday October 19, 2015: 
The Flow Lab is now using iLab for scheduling appointments to use equipment. You should have received an email with detailed instructions for activiating your account in iLab and how to reserve equipment. All future appointments scheduled using Appointment Quest will be transferred to iLab. Contact Dagna Sheerar with any questions or concerns.

 

Once you have attended the lecture series make an iLab appointment for two hours on:


BD FACSCalibur (7016 WIMR) - Tuesday 12-3pm 
BD LSRII - Wednesday 12-3pm

BD LSR Fortessa - Thursdays, 12-3pm

MACSQuant10 - Thursdays, 12-3pm

ImageStream - Monday 12-3pm)

 

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Training

The two-hour hands-on training is intended for those with little or no flow cytometry experience. It will teach the basics of instrument set-up and data collection. Fluorescent Beads will be used for training. Please do not bring cell samples.

 

Once the two-hour training is completed or for those who have some experience operating a flow cytometer, it will be necessary to schedule an hour of assisted time with user samples to discuss specific assay data collection. No user will be allowed to operate the cytometers independently until these training sessions are completed.

 

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Analysis Computers

Offline data analysis computers are available to laboratory users only and are located at both the 7th floor WIMR location and at the McArdle laboratory. Computers are loaded with a variety of analysis packages including FlowJo, CellQuestPro, FCS Express, ModFit, WinList, and Image analysis, as well as standard Microsoft Office software.

There is no charge for the use of these computers. However, they must be used exclusively for the analysis of data generated at this laboratory. Computers are available on a first-come, first-served walkup basis for users of the facility. 

 

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Benchtop Analyzers

The benchtop instruments used for this purpose are the FACSCalibur (488nm and 633nm lasers),  LSRII (488nm, 405nm, 561nm, 355nm, and 642nm lasers), Fortessa (488nm, 403nm, 561nm, 355nm, and 643nm lasers with High-Throughput Sampler for automated sampling of 96 and 384 well plates) and MACSQuant 10 (488nm, 405nm, and 633nm lasers with automated tube and 96 well plate sampling option and in-line magnetic bead separation column option). 

 

New User Training: Schedule two hours. Those with little flow cytometry experience: You MUST attend the Introduction to Flow Cytometry lecture series prior to signing up. Assisted appointments: For those who need assistance operating benchtop instruments for data collection.


When signing up for appointments, insure all fluorochromes to be used can be excited with the lasers listed. The instruments are available for self-run appointments for which no assistance is provided or for staff-assisted appointments for those who need assistance in data collection or instrument operation. Staff-assisted appointments can be used by those who have not received any instrument training but need samples analyzed, as well as by those who need assay-specific instruction.

 

Instrument Maps:

Use iLab to make an appointment for:

 

BD FACSCalibur (7016 WIMR)

All users operating cytometers after staff hours (8pm-6am) are responsible for shutting down the cytometer after use. Please contact staff with any questions.

 

BD FACSCalibur (118 McArdle) 


BD LSRII

All users operating cytometers after staff hours (8pm-6am) are responsible for shutting down the cytometer after use. Please contact staff with any questions.

 

BD LSR Fortessa

All users operating cytometers after staff hours (8pm-6am) are responsible for shutting down the cytometer after use. Please contact staff with any questions.

 

MACSQuant10 

The MACSQuant is a benchtop cytometer is with tube and plate loaders, integrated MACS magnetic bead separation column. 488nm has four detectors, 405 nm has two detectors and 633 nm has two detectors. Trained users only.

 

ImageStream

The ImageStream MarkII is an imaging cytometer for cells in suspension. It has high resolution CCD cameras instead of Photomultiplier Tubes for detection and is equipped with 20x, 40x and 60x objectives, 6 detection channels and 2 lasers (488nm and 642nm). 

 

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Sorters

The facility has two high-speed sorters: one located in the McArdle laboratory and one at the 7th floor WIMR location. Each sorter has some unique capabilities for the maximum applications, but they both can sort up to four separate populations simultaneously. Operated by staff ONLY. They are both housed under BSL2 conditions. Note: Samples exceeding BSL2 levels when aerosolized cannot be sorted.

 

Sorter Maps:

Use iLab to make an appointment for:


BD FACSAria without Biosafety Cabinet "Jill” 

The SORP BD FACSAria is a digital system equipped with five lasers (UV, 405 nm, 488 nm, 532 nm and 640 nm). The Aria can simultaneously collect fluorescent emissions from 16 different fluorophores.  Please contact Flow Lab staff us if you have questions regarding fluorophore compatibility. 

 

BD FACSAria with Biosafety Cabinet "Jack” 

The FACS AriaIII is a cell sorter equipped with four lasers (403 nm, 488 nm, 561 nm and 637 nm). This sorter is contained in a custom Biological Safety Cabinet. Sorts containing human cells, retroviral transfections or as determined in your lab's Biological Safety Protocol MUST be sorted on this instrument.

 

Adding Flow Cytometry to your Bio ARROW Protocol 

UW Flow Lag Hood Certification Dates

 

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Microscopes

Please consult with the staff before scheduling an appointment if you have not used the microscope before. 

 

Use iLab to make an appointment for:

 

Olympus Microscope 

The Olympus BX51 Fluorescent Microscope is equipped with CCD camera, SPOT imaging software, 10x, 20x and 40x objectives and the following excitation/emission filters:  480/20 ex/510LP em, 535/30 ex/580LP em, 365/10 ex/420LP em, 535/50 ex/346/75 em, and brightfield.   

 

Nexcelom Cellometer

The Nexcelom Cellometer Auto T4 Cell Viability Counter 

 is a PC-based instrument for one-step 4x bright field image capture and analysis. This state-of-the-art cell counter performs count, size, and Trypan Blue viability analyses that are 10x faster and more accurate than manual counting. This instrument is ideal for cell lines and purified primary cells, including MCF-7 breast cancer cells, PC3 prostate cancer cells, and 1600 other cell types, due to its advanced software that assesses clumpy cells and omits debris.

 

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FACSCaliburFirtessaFortessa MACSQuant ImageStream MarkII

AriaIIFACS AriaIII