THE UNIVERSITY OF NEWCASTLESCHOOL OF BIOMEDICAL SCIENCES & PHARMACY STANDARD OPERATING PROCEDURE

PROCEDURE NO: MOD: Page:

Procedure Type:

General Equipment Procedure

Title:

Invitrogen Countess- Automated Cell Counter

1.

GEP 094 1st Issue 1 of 5

Risk Assessment: This Risk Assessment is to be used as a general guide and as such, cannot accommodate all the varying factors that may be encountered when using this equipment. Therefore, personnel are requested to conduct their own Risk Assessment before using this equipment to include any extra hazards introduced by the task performed.

TASK PERFORMED Using the Invitrogen Countess Automated Cell Counter o The countess accurately determines cell concentrations between 1 x 104 to 1 x 107 cells per ml.

HAZARDS 1. Sharps- pipette tips 2. Trypan Blue: R36- Irritating to eyes R38- Irritating to skin R45- May cause cancer 3. Hazard Electricity

RISK ASSESSMENT 1. Sharps- Low risk if disposed of correctly 2. Trypan Blue- Low Risk if correct PPE is worn and handled correctly 3. Electricity- Low risk if equipment is maintained properly and inspected before use.

RISK CONTROL WRITTEN BY

CHECKED BY

AUTHORISED BY

Emma Beckett

Linda Howland

Philip Hansbro

10/11/11

14/11/11

15/11/11

NAME (signed) NAME (printed) DATE

Distributed To:

GEP Master File / GEP Lab File

REVIEWED BY

THE UNIVERSITY OF NEWCASTLESCHOOL OF BIOMEDICAL SCIENCES & PHARMACY STANDARD OPERATING PROCEDURE Procedure Type:

PROCEDURE NO: MOD: Page:

GEP 094 1st Issue 2 of 5

General Equipment Procedure

Title: 1. Wear PPE- Lab Coat, Gloves, Closed in Shoes, Safety Glasses. 2. Check electrical power cord is in good condition prior to use. 3. Make sure area is free of water before switching on machine. 4. Dispose of sharps immediately in an appropriate sharps container. 5. Trypan Blue- S24/25 - Avoid contact with skin and eyes S36/37/39 - Wear suitable protective clothing, gloves and eye/face protection

2.

Calibration: 2.1.

Frequency- Machine is supplied pre-calibrated- recalibration is only needed if you

use a trypan blue other than supplied or upgrade the firmware. Note the countess contains delicate optics- move and handle with care

3.

4.

2.2.

Method

2.3.

Records

Monitoring: 3.1.

Frequency

3.2.

Method

3.3.

Records

General Operation: 

Equipment: o Invitrogen Countess automated cell counter o p20 pipette o Countess USB drive



Materials: o 0.4% trypan blue (Invitrogen) o Pipette tips o Counting Chamber Slides (Invitrogen)



Method: o

Turn on Countess- the following start up screen will display

THE UNIVERSITY OF NEWCASTLESCHOOL OF BIOMEDICAL SCIENCES & PHARMACY STANDARD OPERATING PROCEDURE Procedure Type:

PROCEDURE NO: MOD: Page:

GEP 094 1st Issue 3 of 5

General Equipment Procedure

Title:

o

Load Sample: Add 10ul sample to 10ul of 0.4% trypan blue stain in a separate tube or plate well.

o

Add 10ul of this mixture to the half moon shaped chamber port (marked A or B). You may load 1 or both chambers (for best results always handle slides by the edges). If your samples are infectious or potential infectious, the countess can be moved to the BSCII, move gently and clean before removing.

o

Insert Slide: sample A end first into the slide port in the middle of the instrument. If inserted correctly you will hear a soft click.

o

o

Optimize Image: Press Zoom and use the focus knob and adjust image (live cells should have bright centres and dark edges, dead cells should have uniform dark blue colour throughout as they have taken up the

THE UNIVERSITY OF NEWCASTLESCHOOL OF BIOMEDICAL SCIENCES & PHARMACY STANDARD OPERATING PROCEDURE Procedure Type:

PROCEDURE NO: MOD: Page:

GEP 094 1st Issue 4 of 5

General Equipment Procedure

Title: trypan blue stain. Use the grid to navigate around the slide to check multiple fields. Note: After you have counted the first sample, you may not have to use the Focus knob again. If measuring multiple samples of the same approximate size, you may lock the Focus knob. You are able to unlock the knob later, to adjust the image, if needed o

Press “Count Cells” on screen to obtain result.

o

After approx 30 second count (total, live, dead will be displayed on the screen. Either record the result manually, or insert supplied USB drive and press save.

o

You can use the Details button to see how each cell was counted. Objects are circled in blue, (live) red (dead), or black (excluded). See more details on the data as well as graphical representation of the data, press the More Data button.

o

To count the second chamber: remove the slide by gently pressing in to eject it, then pulling out. Turn slide around, re-insert and repeat the procedure.

o

The Countess memory holds one set of data. To save your data for future analysis or archiving, you must record the data or save after each reading.

o

After recording or saving the data, remove and discard the entire slide appropriately as biohazardous waste. Ie. Use a new slide for each sample/pair of samples.

o

See manual for instructions for saving data to the supplied USB- please return USB to the Countess after use.

o 5.

Turn off the machine after use.

Cleaning: 5.1.

Frequency: As required

5.2.

Method : Clean the surface of the Countess™ instrument with a damp cloth. To

clean the LCD screen, turn off the Countess™ instrument, disconnect the power cable, and clean the LCD screen with a soft cloth lightly moistened with LCD cleansing detergent. Cleaning the screen with excessive force can damage the LCD the screen. Wipe the screen dry immediately. Do not reuse the counting chambers. 6.

Maintenance:

THE UNIVERSITY OF NEWCASTLESCHOOL OF BIOMEDICAL SCIENCES & PHARMACY STANDARD OPERATING PROCEDURE Procedure Type:

PROCEDURE NO: MOD: Page:

GEP 094 1st Issue 5 of 5

General Equipment Procedure

Title: 6.1.

Frequency & Method: The Countess™ automated cell counter does not need

regular maintenance. To troubleshoot problems with Countess™ cell counter, contact Technical Support (page 26). Do not perform any repairs or service on the Countess™ instrument to avoid damaging the instrument. 6.2. 7.

Instruction Manual: 7.1.

8.

Records

Available for download at www.invitrogen.com

Waste Disposal: Slides should be disposed of in appropriate biohazard sharps container. NB: University, MSDS and other regulatory guidelines must be adhered and referred to.

9.

Repairs: Contact Laboratory manager and Invitrogen

10.

Attachments: 10.1.

11.

Instruction Manual- is available for download at www.invitrogen.com

References: 13.1. http://www.invitrogen.com/etc/medialib/en/filelibrary/CellCulture/PDFs.Par.13233.File.dat/Countess_Manual_0909.pdf

12.

Review History: 12.1.

Issue Number: Date Issued:

12.2.

Issue Number: Date Issued: Reason for Change:

1st Issue