ClonePix 2 System

High-throughput screening and objective selection of mammalian cells

Automatically screen more clones in less time than conventional techniques, select cells with optimal expression levels, and pick colonies with accuracy with the ClonePix™ 2 System. ClonePix Systems are now used in over 100 laboratories around the world to increase workflow productivity, leaving more time to better characterize target proteins and run new projects. A large number of biopharma companies have implemented ClonePix Systems in their routine use and data are cited increasingly in scientific publications and conferences

  • Select cells with optimal expression levels with improved ranking consistency. Increase probability of finding optimal producers and select cells 
  • Cut cell line/antibody development times – avoid limiting dilution
  • Reject poor performers at an early stage based on expression levels in situ
  • Pick colonies with accuracy and confidence. Reduce risk of colony disturbance with robotics redesign.
  • Improve workflow efficiency with fewer plate handling steps

Antibody Development for Biotherapeutics

Select optimally secreting cell lines

Select Clones Secreting Optimal Monoclonal Antibodies

  • Screened 5463 colonies (5 plates) using white light and fluorescent imaging
  • Selected 16 high secretors
  • CHO-S cells grown in CloneMedia-CHO
  • Screened for IgG secretion by addition of CloneDetect agent
  • Clones ranked according to fluorescence
    • User defined selection parameters
  • System accurately picks clones for expansion
    • Proximity settings avoid picking neigboring clones
    • System transfers each colony into one well of a 96-well microplate 
CHO-S expressing human IgG: stiched image
of PetriWell-1 plate picked on day 13
Ranking plot - Clones ordered by
fluorescence intensity (secretion)

Imaging Principle - Label-Free Detection of Secreted Antibody


  • Secreting clones grown in semi-solid media
  • CloneDetect agent added
  • Secreted antibodies form fluorescent precipitate

Development of cell lines for biotherapeutics

Select Optimal Producer NS/0 and CHO Cell Lines Using ClonePix System

  • Screens more clones than traditional methods, increasing the probability of finding rare, high secretors
  • Automatically isolates clonal colonies, removing the need for limiting dilutions
  • In situ indication of high titer cell lines removes unwanted clones from further processes


Comparison of Techniques:


ClonePix System

  • From parental cell line to shake flask: 1 month
  • 10000 clones screened

Limiting dilution

  • From parental cell line to shake flask: 2 months
  • 1000 clones screened
Comparison of Cloning Methods:


ClonePix System

  • 2500 clones per week
  • 1 week cloning

Limiting dilution

  • 300 clones per week
  • 3 weeks cloning

Comparison of Top Clones from ClonePix System and Limited Dilution

Clone Titer (g/L) qP (pcd)  
Clone 1 4.5 54.6 ClonePix System
Clone 2 4.4 44.6
Clone 3 4.3 49.4
Clone 4 4.0 43.8
Clone A 2.9 32.7 Limiting Dilution
Clone B 2.8 21.0
Clone C 2.7 20.9
Clone D 2.6 29.0
  • Cell lines selected were about 2- fold more productive compared to those selected by conventional approach
  • Sub-clones from same parent (NS/0 cells)

High Titers of Final Clones from ClonePix System Method

Clone Titer (g/L)
1 5.8
2 5.3
3 5.7
4 5.8
  • CHO cells - 14 day fed batch process in shake flasks
  • High titer clones obtained (NS/0: 4-5 g/L, CHO: 5-6 g/L) even prior to process optimization
Conclusion from MedImmune: ‘ClonePix System is a powerful tool in cell line development. This method makes selecting the optimal producers faster and less labour intensive and shortens cell line development time.’

Further Information:

Rapid and efficient selection of high producing mammalian cells secreting therapeutic proteins/peptides: Application Note Download

Eliminate unstable clones

  • Reveal clonal instability with ClonePix systems
  • Re-plate aliquots of selected clones into semi-solid media and, within 4-7 days, re-image to verify and compare production rates of the daughter clones and/or re-screen for sub-clones within 7-14 days. 

Example shows results from a selection of stable clones by second round screening System transfers each colony into one well of a 96-well microplate

verify clonal stability, cell line stabilization, clonal screening
Top 2% of transfected population
of suspension- adapted CHO cells
collected and assayed for productivity
Productivity vs. fluorescence after second
round screening. Note elimination of
high fluorescence, low productivity
(unstable) clones

Cell confluence and cell number can be tracked by CloneSelect Imager

Antibody Discovery for Research

Identify higher-titer hybridoma cell lines for antibody development

  • Parental hybridoma cell line was expanded and screened on ClonePix system
  • High yield clones picked by ClonePix System and monitored for growth on CloneSelect Imager System
  • Top growers screened for antigen specificity by target peptide binding assay
  • High valued, specific sub-clones were isolated by pilot production and cell banking
  • Workflow reduced hybridoma screening time by >50% finding optimal producers secreting IgG at higher titers
White light image
Fluorescent image

High-throughput screening of hundreds of sub-cloned colonies from parental hybridoma material on ClonePix system aids rescue and stabilization of a high-titer hybridoma cell line secreting highly specific monoclonal antibody to an immunogenic viral antigen.

Further Information

Enhanced Development of Virus-Specific Hybridomas using ClonePix and CloneSelect Imager Technologies: Application Note Download

Imaging Principal - Label-Free Detection of Secreted Antigen-Specific mAbsimage_antigen_0.gif

  • Hybridoma clones grown in semi-solid media
  • Complex Initiation Factor (CIF) traps secreted mAb
  • Fluorescently conjugated antigen targets IgG-secreting clones

Further Information

Screen and select antigen-specific hybridomas or B Cells

Screening Antigen-Specific Hybridomas

  • Screen and select antigen-specific clones in situ
  • Suitable for a broad range of antigens
    • - 160kD multimeric protein to 2.6kD phosphopeptide
White light image
FITC conjugated 60kD antigen-specific
IgG producing clone
White light image
FITC conjugated 60kD control protein

Imaging Principal - Label-Free Detection of Secreted Antigen-Specific mAbsimage_antigen_0.gif

  • Hybridoma clones grown in semi-solid media
  • Complex Initiation Factor (CIF) traps secreted mAb
  • Fluorescently conjugated antigen targets IgG-secreting clones

Further Information

Protein Expression and Production

Selection and Development of GPCR Expressing Cell Lines

Detection and Selection of Cells Expressing GPCR at Low Endogenous Levels

  • Positive (CHO-M1) and negative (CHO-K1) cells plated in CloneMedia and incubated for 8-10 days
  • Labeled antibodies added and cells imaged in brightfield and fluorescent channels
  • Cells visible in both channels picked and cultured
  • GPCR expression validated by fluorescent calcium signals on FLIPR Tetra System
System locates and identifies
clones using brightfield
Cells positive for GPCR expression
are shown in fluorescent channel

Further Information

  • Rapid Selection and Development of GPCR Expressing Mammalian Cell Lines Using Novel ClonePix Technology: Application Note Download

Select by intrinsic reporter expression

Example: Detection of Intrinsic GFP Reporter 

  • Human breast cancer cells (MCF-7) transfected with internally expressed GFP-fusion protein
  • Cells grown as adherent colonies in MEM Earle’s liquid medium + 10% FBS
  • Day 8: Clones expressing highest GFP selectively picked
White light
After picking

Select by cell surface expression marker

  • Adherent CHO K-1 stably transfected with P2Y1 receptorsurface_expression_0.jpg
  • Cells grown in CloneMatrix based semi-solid medium
  • Day 8: anti-P2Y1 receptor polyclonal conjugated with FMAT-Blue added by atomizer
  • Day 9: High expressers of P2Y1 receptor screened and picked

Further Information:

Rapid Automated Selection of Mammalian Cell Colonies by Cell Surface Protein Expression: Application Note Download

Select Secretors of a tagged recombinant Protein

Detection of Secreted Monomeric His/FLAG-Tagged Protein

  • Adherent CHO cells stably transfected
  • Grown as suspended colonies in CloneMatrix-based semi-solid medium with FITC-conjugated anti-His and anti-FLAG polyclonals
  • High secretors screened and picked at day 8
White light image
FITC image

Imaging Principal


Stem Cell Screening/Maintenance of Stem Cell Lines

Maintain lines utilizing colony Morphology (white light)

Maintenance of Human Stem Cells (BG01V hES) Utilizing White Light and Morphology

  • Human stem cells (BG01V hES) grown as colonies on feeder layer in liquid media
  • Plates imaged and colonies picked based on morphology
Stem cell clones shown by
white light imaging on
ClonePix System
Typical picked clone collected
to 96-well plate. Imaged
on CloneSelect Imager

Colony formation

Cells are grown in source plates containing semi-solid CloneMedia. This approach facilitates plating out of large numbers of cells and ensures formation of discrete colonies for subsequent recovery as independent clones.

Colonies of commonly used cell types growing in CloneMedia. Images captured using CloneSelect Imager

CHO colonies, serum-free
in CloneMedia-CHO
imaged on day 8 post-plating.
Hybridoma colonies in
Myeloma imaged on day
8 post-plating.
HEK293 colonies, serum-free,
suspension-adapted, in
CloneMedia-HEK imaged
on day 13 post-plating.
CHOK1SV colonies, serum-free,
in CloneMedia-CHOK1SV
imaged on day 9 post-plating.

Choice of Detection Methods

Detection using white light for imaging and picking, flurescence for indicating expression levels

Detect Recombinant Proteins and Expression Markers

The protein construct contains epitope tag(s)
e.g. His, FLAG® or Fc using tag-specific agents.

Label-Free Detection of Antigen-Specific mAbs from Hybridomas 

Fluorescently-conjugated or tagged antigen plus CIF
(complex initiation factor) are used to generate precipitation.

Label-Free Detection of Secreted Antibodies 

CloneDetect fluorescently-conjugated agents enable in situ
detection of secreted mouse, rat or human IgG.
These agents can be added as liquids or simply
sprayed onto the colonies. Relative secretion
levels are then measured across 1000’s of clones in parallel.

Imaging and Analysis

Colonies are imaged in white light and fluorescence then analyzed by the system.

White light imaging identifies and
measures clone morphology,
size and proximity
Fluorescence imaging identifies
levels of expression

Ranking and Data Display

Ranking plot
  • Clones ranked according to fluorescence
  • User defines selection parameters – system selects clones
    • Morphometric criteria such as size, roundness and proximity to neighbors
    • Quantitative criteria such as protein secretion or specific protein production
  • Bar-coded source and destination plates enable data tracking.
  • Images of all picked clones taken before and after picking are stored with their data, including picking coordinates

Related Collateral

Brochures and Catalog


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Product Support

For product configurations and pricing on ClonePix 2 System, please contact us

Latest Citations:
For a complete list of citations, please click here.


High-throughput ClonePix FL analysis of mAb-expressing clones using the UCOE expression system

JJC Hou, BS Hughes, M Smede, KM Leung, K Levine… - New biotechnology, 2014 - Elsevier
Therapeutic recombinant monoclonal antibodies (mAbs) are commonly produced by high-
expressing, clonal, mammalian cells. Creation of these clones for manufacturing remains 
heavily reliant on stringent selection and gene amplification, which in turn can lead to ...

Rapid isolation of antigen-specific clones from hybridoma fusions

CJ Mann - Nature Methods, 2007 -
... The Genetix ClonePix FL is a powerful way to find and collect antigen-specific clones from
hybridoma fusions in a rapid one-step process—shortening the process timeline and allowing
parallel interrogation of multiple antigens in the monoclonal antibody discovery process. ...

Stem and iPS cell selection: quantitation of surface marker (SSEA1) and intracellular GFP

CJ Mann, ENC Newman, DJ Whitney, NJ Latchem… - Nature …, 2007 -
... The Genetix ClonePix FL technology, already established for the rapid one-step identification
and isolation of hybridomas and cell lines for biopharmaceutical production, can be used to isolate
and collect stem cells expressing specific cell surface and intracellular proteins. ...

Improving Clone Production for Increased Protein Yield from Mammalian Cell Lines

S Williams, R Cranenburgh - Innovations in Pharmaceutical …, 2008 -
... An alternative approach for the screening of secreting clones is the use of cell imaging systems
such CellXpress marketed by Sigma-Aldrich Corporation, and ClonePix FL from Genetix Ltd.
ClonePix FL can screen around ten thousand in three weeks. ...

Rapid automated selection of mammalian cell colonies by cell surface protein expression

ENC Newman, D Whitney - Nature Methods| Application Notes, 2007 -
... Top of page Abstract. Genetix's unique ClonePix FL technology uses our ability to
grow mammalian cells into clonal colonies suspended in semisolid media, and image
these colonies using fluorescence assays and markers. ...

An improved clone selection method

U Jamnikar, M Blas, D Gaser… - Proceedings of the 21st …, 2012 - Springer
... In most cases there are no resources to isolate and analyze more than few 100 clones. ClonePix
FL (CP-FL, Genetix) technology substantially increases the number of screened clones in the
selected pool before expansion in microtiter plates. ... 1.2.1 ClonePix FL. ...

Micropallet arrays for the separation of single, adherent cells

G To'a Salazar, Y Wang, G Young, M Bachman… - Analytical …, 2007 - ACS Publications
... the sample during collection. ClonePix (Genetix, Hampshire, UK) is an automated
system that uses image recognition to guide a suction pipet that aspirates colonies
of loosely adherent cells from plates. The system requires ...

High‐Throughput Technologies in Bioprocess Development

T Carrier, E Heldin, M Ahnfelt, E Brekkan… - Encyclopedia of …, 2010 - Wiley Online Library
... Cell Line Development Technology Developer ClonePix Fl Genetix LEAPTM Cyntellect FACS
Multiple developers ... The ClonePix Fl technology, developed by Genetix, capitalizes on a system
in which mammalian cells are cultured in a semisolid media and then imaged using 1 ...

Screening and Subcloning of High Producer Transfectomas Using Semisolid Media and Automated Colony Picker

S Dharshanan, CS Hung - Monoclonal Antibodies, 2014 - Springer
... 3.2 Sanitization and Calibration of ClonePix FL System. Diligently follow the instructions of the
pre-programmed software in the ClonePix FL system. ... 10. Sanitize the system with ultraviolet light
for 10 min. 3.3 Selection of High-Producing Transfectomas Using ClonePix FL System. ...

Screening and Selection of GPCR-Expressing Cell Lines

A Glaser - 2014 -
... The ClonePix™ 2 System from Molecular Devices represents a proven, one-step method of
screening large cell populations rapidly. ... The ClonePix 2 System is a platform enabling the
identification and isolation of GPCR clones in an accurate and rapid manner. ...