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Antibody Fixation Considerations

eBioscience Antibody Clone Performance Following Fixation / Permeabilization

It is sometimes possible to stain cells after fixation but this is dependent upon the effects of fixation on the epitope of your protein-of-interest.  The list below contains information about the performance of different clones after fixation using our eBioscience Intracellular Fixation & Permeabilization Buffer Set (cat. no. 88-8824) or after IC Fixation Buffer (cat. no. 00-8222) followed by treatment with methanol. Please note that staining after fixation may allow for intracellular stores of protein to also be stained with the antibody.

Antibody Clone Performance Following Fixation / Permeabilization
AntigenCloneLive cells (Before IC Fixation)1After IC Fixation and Perm Wash1After IC Fixation/ Methanol2
HUMAN        
CD3 OKT3   +++ ++  +
  UCHT1  +++ +++  +++
  SK7  +++ ++*  +++
CD4 OKT4  +++ +*  +/-
  RPA-T4  +++ +++*  +
  SK3  +++  +++  +
CD5 UCHT2  +++ +++  +++
CD8 OKT8  +++ +/-   - 
  RPA-T8  +++ +++  +/-
  SK1  +++ +++*  +/-
CD8b SIDI8BEE  +++ ++  ++
CD11a HI111  +++ +++  +
CD11b CBRM1/5  +  -  +++
  ICRF44  ++  -  ++
CD11c 3.9  ++   +*  ++
CD14 61D3  +++  +++*  ++
CD19 HIB19  +++  ++*  +/-
CD20 2H7  +++  +++  -
CD25 BC96  +  -  +/-
CD27
O323
 +++   +*  +
CD31 WM59  +++  ++*  +++
CD33 p67.6  +++  +++*  +++
CD38 HB7  ++  -  ++
  HIT2  ++  -  ++
CD40
5C3
 ++  -  ++
CD44 IM7  +++  +++  +++
CD45 2D1  +++  +++  +++
  HI30  +++  +++  +++
CD45RA HI100  +++  +++  +++
CD45RO UCHL1  +++  +++  ++
CD56 MEM-188  +  -  +/-
  CB56  +  -  +/-
  CMSSB  ++
 +/-*  -
CD57 TB01  ++  ++  ++
CD62L DREG-56  ++  -  ++
CD69 FN50
 +++  ++  ++
CD80 2D10.4  +  -  +
CD83 HB15e  ++  ++  ++
CD86 IT2.2  +  +*  +
CD94 HP-3D9  ++  ++  ++
CD95 DX2  +  -  +
CD127 RDR5  ++  +*  ++
CD161 HP-3G10  ++  ++*  +
MOUSE        
CD3 145-2C11 ++  + +/-
  500A2 +++  +++* +++
  17A2 +++  +++ +++
CD4 GK1.5 +++  ++* +++
  RM4-5 +++  +++* +++
CD8 53-6.7 +++  ++ ++
CD11b M1/70 ++  ++ ++
CD11c N418 +++  +++ ++
CD19 1D3 +++  ++* -
  MB19-1 ++  - -
CD24 M1/69 +++  +++ ++
CD25 PC61.5 +  - +
  3C7 ++  - +
  7D4 ++  ++ +
CD39 24DMS1 +  - ++
CD44 IM7 ++  ++* ++
CD45 30-F11 +++  +++ +++
CD45R(B220) RA3-6B2 +++  ++ +++
CD49b DX5 ++  - +
CD69 H1.2F3 ++  - -
CD185(CXCR5) SPRCL5 ++  - -
GR-1 RB6-8C5 +++ +++ ++
NK1.1 PK136
++ + +
IgD 11-26c +++
+++*
++
IgM ll/41 ++ ++* ++
TCR beta H57-597 ++ ++* ++
RAT        
CD25 OX39  +  -  
CANINE        
CD4 ykix302.9 +++  +  
CD5 ykix322.3 +++  -  
CD8a ycate55.9 +++  ++  
CD44 ykix337.8 +++  ++  
CD45 ykix716.13 +++  ++  
CD45R ykix753.22 +++  ++  
CD90 ykix337.217 +++  ++  
MHC Class II ykix334.2 +++  +++  

 

1 Testing carried out with eBioscience Fixation & Permeabilization Kit for 15-30 minutes at room temperature followed by a Perm wash, see Best Protocols here  (Protocol A).  

*Please note the clones with * indicate longer fixations are detrimental to staining.

2 Testing carried out with 15-30 minutes at room temperature in 50% IC Fixation followed by 30 minutes 90-100% Methanol incubation on ice, see Best Protocols here (Protocol C).