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Table 1 Overview of the marker panels of primary monoclonal antibodies and 7-AAD to immunophenotype equine MSC

From: Characterization and profiling of immunomodulatory genes of equine mesenchymal stromal cells from non-invasive sources

 

Subset

Marker

Company

Clone

Sec Ab

Dilution

Positive control

Multicolor FCM

1

CD29-Alexa488+

Biolegend

TS2/16

 

1:20

MNCs

MHC-II−

Serotec

CVS20

RPE

1:50

MNCs

7-AAD−

Calbiochem

    

CD44-APC+

BD

IM7

 

1:20

MNCs

2

CD105-RPE+

Serotec

SN6

 

1:10

Endothelial cells

7-AAD−

Calbiochem

    

CD90+

VMRD

DH24A

Alexa647

1:100

MNCs

3

CD45-Alexa488−

Serotec

F10-89-4

 

1:5

MNCs

CD73+

Abcam

10f1

RPE

1:5

Lymphocytes

7-AAD−

Calbiochem

    

4

Monocyte-Alexa488−

Serotec

MAC387

 

1:2.5

MNCs

CD79α-Alexa647−

Serotec

HM57

 

1:2.5

MNCs

Single-color FCM

 

MHC-I+

VMRD

PT85A

RPE

1:66

MNCs

Secondary Ab

1 and 3

Sheep anti-mouse IgG-RPE

Sigma-Aldrich

  

1:20

 
 

2

Goat anti-mouse IgG-Alexa647

Invitrogen Corporation

  

1:200

 

Isotype controls

1 and 4

Mouse IgG1-Alexa488

Biolegend

  

1:20

 
 

1-3

Mouse IgG1-RPE

Biolegend

  

1:10

 
 

1

Rat IgG2b-APC

Biolegend

  

1:20

 
 

2

Mouse IgM

BD

 

Alexa647

1:50

 
 

3

Mouse IgG2-Alexa488

Biolegend

  

1:20

 
 

4

Mouse IgG1-Alexa647

Biolegend

  

1:100

 
  1. 7-AAD, 7-aminoactinomycin D; FCM, fluorescence correlation microscopy; MHC, major histocompatibility complex; MNC, mononuclear cell; Sec Ab, secondary antibody.