Thermo Scientific gibco CTS Rotea User manual

For Research Use or Manufacturing of Cell, Gene, or Tissue- Based
Products.
CTS™Rotea™Counterflow Centrifugation
System Process Design
USER GUIDE
Catalog Number A44769
Publication Number MAN0018754
Revision A.0

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The information in this guide is subject to change without notice.
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ARISING FROM THIS DOCUMENT, INCLUDING YOUR USE OF IT.
Revision history: Pub. No. MAN0018754
Revision Date Description
A.0 20 October 2020 New document. User guide for process design workflow.
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©2020 Thermo Fisher Scientific Inc. All rights reserved.

Contents
■CHAPTER 1 Product information .................................................. 7
Product description ............................................................. 7
System overview ................................................................ 8
CTS™Rotea™Counterflow Centrifugation Instrument ............................ 8
CTS™Rotea™Single-Use Kit ................................................. 9
CTS™Rotea™Graphical User Interface (GUI) .................................. 10
Rotea Protocol Builder ..................................................... 11
■CHAPTER 2 Software installation ................................................ 12
Software installation ............................................................ 12
■CHAPTER 3 Rotea Protocol Builder Application ................................ 17
Create or open a protocol ....................................................... 17
Define Kit ..................................................................... 18
CTS™Rotea™Single-Use Kit diagram ........................................ 19
Cell Type ................................................................. 21
Description ............................................................... 21
Kit Type .................................................................. 22
Long Description .......................................................... 22
Step view ..................................................................... 23
Define fluid path and direction of flow ........................................ 24
Step Details .............................................................. 25
Step Type ................................................................ 27
Trigger ........................................................................ 32
Volume ................................................................... 33
Timer .................................................................... 35
Pressure 1 and Pressure 2 ................................................. 36
Bubble Sensor Input and Bubble Sensor Output .............................. 37
Data Entry .................................................................... 38
CTS™Rotea™Counterflow Centrifugation System Process Design User Guide 3

List view ...................................................................... 39
Step summary ............................................................. 40
Edit using List view ........................................................ 42
Loops ........................................................................ 42
Repeat Count ............................................................. 43
Bubble Sensor Not Triggered ............................................... 44
Process Counter .......................................................... 44
Timer (Seconds) and Timer (Minutes) ........................................ 45
Skip Button Triggered ..................................................... 45
Editing loops .............................................................. 46
■CHAPTER 4 Protocol Building Blocks ........................................... 47
Priming ....................................................................... 47
Overview ................................................................. 47
Bubbles .................................................................. 47
Priming volumes ........................................................... 48
Basic priming steps ........................................................ 49
Prime remote sources ...................................................... 52
Manage air in the system ................................................... 53
Establish a fluidized bed ........................................................ 53
Overview ................................................................. 53
The seven parameters for establishing a fluidized bed .......................... 55
Optimize G force and flow rate .............................................. 56
Cell size (d)............................................................... 56
Media density (ρ) .......................................................... 57
Cell concentration ......................................................... 57
Establish a bed in recirculation .............................................. 57
Establish a bed without recirculation ......................................... 59
What a good bed looks like ................................................. 60
Establish a bed: Troubleshooting ............................................ 62
Cell loading ................................................................... 64
Overview ................................................................. 64
Cells loaded into the CFC Chamber .......................................... 64
Bed disruption during loading ............................................... 65
Cell separation during loading ............................................... 65
Re-use input bags ......................................................... 66
Refill the Bubble Trap ...................................................... 69
Connect to bioreactors ..................................................... 70
Wash cells and buer exchange ................................................. 71
Overview ................................................................. 71
Media selection ........................................................... 72
Wash to similar density media ............................................... 72
Wash to more dense media ................................................. 72
Wash to lower density media ................................................ 73
Defensive washing strategies ................................................ 73
Contents
4CTS™Rotea™Counterflow Centrifugation System Process Design User Guide

Elutriation ..................................................................... 77
Overview ................................................................. 77
Improve cell viability ....................................................... 79
Leukopak processing ...................................................... 79
Typical leukaphoresis composition ........................................... 80
Lentivirus clarification ...................................................... 81
Separate cells from microcarriers ............................................ 81
Elutriation techniques ...................................................... 82
Concentrate Recovery ......................................................... 85
Overview ................................................................. 85
Normal step ............................................................... 86
Speed settings ............................................................ 86
Pause step ................................................................ 87
Harvest step .............................................................. 88
Harvest using OD Sensor ................................................... 90
Multi-bite processing ........................................................... 92
Overview ................................................................. 92
Determine the number of cells per bite ....................................... 92
Maximize throughput ....................................................... 93
Process integrity ............................................................... 94
Overview ................................................................. 94
CTS™Rotea™Single-Use Kit ................................................ 94
Check manual clamps ...................................................... 95
Verify reagent connected and not clamped ................................... 95
Check for closed clamps ................................................... 95
■CHAPTER 5 Rotea™Process Model ............................................. 96
Counter flow centrifugation forces ................................................ 96
Predict process settings ........................................................ 98
Select and format media .................................................. 100
Create custom media ..................................................... 102
Model particle behavior ................................................... 103
Change and delete particles from a model .................................. 106
Create custom particles ................................................... 107
Edit custom particles and custom media .................................... 107
■CHAPTER 6 Rotea™Simulation software ...................................... 109
Rotea™Simulation software .................................................... 109
■CHAPTER 7 Design new protocols ............................................. 115
Plan the kit configuration ....................................................... 115
Plan the protocol ............................................................. 115
Prepare for the first trial run .................................................... 116
Contents
CTS™Rotea™Counterflow Centrifugation System Process Design User Guide 5

Water runs ................................................................... 116
First run with cells ............................................................. 116
Analyze the outcome .......................................................... 116
Refine the protocol to achieve target cell outcomes ............................... 116
Add process integrity features .................................................. 117
■APPENDIX A Safety .............................................................. 118
Symbols on this instrument .................................................... 118
Standard safety symbols .................................................. 119
Control and connection symbols ........................................... 119
Conformity symbols ...................................................... 119
Safety information for instruments not manufactured by Thermo Fisher Scientific ..... 121
Instrument safety ............................................................. 121
General ................................................................. 121
Physical injury ............................................................ 122
Electrical safety .......................................................... 122
Cleaning and decontamination ............................................. 123
Safety and electromagnetic compatibility (EMC) standards ......................... 123
Safety standards ......................................................... 123
EMC standards ........................................................... 124
Environmental design standards ............................................ 124
Chemical safety .............................................................. 125
Biological hazard safety ....................................................... 126
■APPENDIX B Documentation and support .................................... 127
Customer and technical support ................................................ 127
Limited product warranty ...................................................... 127
Contents
6CTS™Rotea™Counterflow Centrifugation System Process Design User Guide

Product information
IMPORTANT! Before using this product, read and understand the information in the “Safety” appendix
in this document.
Product description
The Gibco™CTS™Rotea™Counterflow Centrifugation System is an automated, closed-flow benchtop
instrument that can perform a wide variety of cell processing steps across dierent cell types.
This CTS™Rotea™Counterflow Centrifugation System Process Design User Guide provides the user
with an overview of the Rotea Protocol Builder application and describes some of the commonly used
features and strategies to assist the user with the development of protocols.
Note: The user is responsible for the development, optimization, qualification, storage and
management of protocols used on the CTS™Rotea™Counterflow Centrifugation System.
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CTS™Rotea™Counterflow Centrifugation System Process Design User Guide 7

System overview
CTS™Rotea™Counterflow Centrifugation Instrument
The CTS™Rotea™Counterflow Centrifugation Instrument is an automated benchtop instrument that
incorporates several important features that enable the user to develop and optimize protocols while
ensuring the integrity of the process.
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Figure 1 Instrument – Front View
1Instrument Door
2Pinch Valves (J & K)
3Kit Location Button
42D Barcode Reader
5CFC Chamber Detector
6CFC Chamber Carrier
7Camera
8Moisture Sensor
9Pushbutton Controls
10 Optical Density Sensor & Pressure Sensor (P1)
11 Peristaltic Pump
12 Pump Clamp Arm
13 Pinch Valve (H)
14 Kit Location Button & Pressure Sensor (P2)
15 Door Latch
16 Pinch Valves (A, B, C, D, E, F, & G)
17 Bubble Sensors
18 Hanger Posts
Chapter 1 Product information
System overview
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8CTS™Rotea™Counterflow Centrifugation System Process Design User Guide

CTS™Rotea™Single-Use Kit
The CTS™Rotea™Single-Use Kit can be flexibly configured by the user to perform a range of cell
processing applications such as separation, isolation, buer exchange, wash and concentrate. The
Single-Use Kit comprises eight sterile weldable tubes to which the user connects input and output
bags/vessels to suit their specific process. The Single-Use Kit also includes a 2D Barcode for kit
identification and labelling to aid kit assembly.
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Figure 2 CTS™Rotea™Single-Use Kit
1Valve Hole & Tube ID
2Carrier Frame
3Bubble Trap
4Location Hole
5CFC Chamber
6Rotary Coupling
72D Barcode
8Pump Tubing
9Tube Retainer
10 Valve H Hole
11 Location Hole
12 Kit Tubing (Input and Output)
Chapter 1 Product information
System overview 1
CTS™Rotea™Counterflow Centrifugation System Process Design User Guide 9

CTS™Rotea™Graphical User Interface (GUI)
Common Callouts and Arrows
1. Copy-paste a callout or arrow to use in this SVG.
Note: If you need more advanced callouts or arrows
use the TechComm_Inkscape_Callout&Arrow_Libary.
3. Delete this text, this rectangle, and unused
callouts, arrows, or other SVG elements
before adding this SVG to the repository.
2. Edit number and/or line-length, as needed.
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The CTS™Rotea™Graphical User Interface (GUI) enables users to download and run protocols on the instrument. The
Rotea™GUI also includes the ability to make in-process adjustments to the G force and flow rate and has an integrated
high-resolution color camera to visualize the behavior of cells in the CFC Chamber in real-time. This provides the user with
an interactive environment for protocol development and optimization.
Chapter 1 Product information
System overview
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10 CTS™Rotea™Counterflow Centrifugation System Process Design User Guide