EuroMAbNet: European Monoclonal Antibodies NetworkEuroMAbNet. European Monoclonal Antibodies Network homepage

Menu

Antibody Identity card

Antibody information [CD85a (LILRB3/ILT5)]

FRAS92B

  • Clone name FRAS92B
  • Description Rat monoclonal
  • Antigen used Human HIS-GST-CD85A fragment (463-631aa)
  • Epitope Unknown
  • Isotype IgG1
  • Confirmed species reactivity Human
  • Ab used Rat mAb CNIO (tissue culture supernatant)
APPLICATIONRecommended concentrationStatusProtocol
Western blotting (WB)Neat tissue culture supernatantWorkingWestern Blotting (WB)
Immunocytochemistry (ICC)1:3 tissue culture supernatantWorkingICC in frozen tissue and cytospin preparation
Immunohistochemistry (IHC-P)1:3 tissue culture supernatantWorkingBOND-MAX automated immunohistochemistry
Immunoflourescence (IF)1:3 tissue culture supernatantWorkingImmunofluorescence staining
Flow cytometry (FC)Not tested
IHC-P SpeciesNot tested

Ab ID Card application validation / characterisation

WB Validation
Over-expression/cross reactivity
CD85a (LILRB3/ILT5) antibody WB Validation: Over-expression/cross reactivity
To confirm the lack of cross-reactivity with the other CD85 family members, western blotting of the FRAS92B mAb was performed using cell extracts of HEK239T transfected with all the CD85 family members (CD85A-K). A specific band of 80kDa was only detected in the HEK-CD85A cell extracts. Anti-GAPDH was used as loading control.

Show image in a new window

Gene inactivation
Not tested
WB Characterisation
Endogenous expression
CD85a (LILRB3/ILT5) antibody WB Characterisation: Endogenous expression
The detection of CD85a endogenous expression using the FRAS92B mAb was confirmed by Western blotting using the myeloid-derived MUTZ3 cell line (widely used as a model for dendritic cells) and the myeloma-derived U266 (negative control). A specific band of 80 kDa was detected in MUTZ3 cell extract while no expression was found in U266. Anti-actin was used as loading control.

Show image in a new window

ICC / ICC-P Validation
Over-expression/cross reactivity
CD85a (LILRB3/ILT5) antibody ICC Validation: Over-expression/cross reactivity
The FRAS92B mAb recognised human CD85A protein by ICC on cytospin preparations of transfectants. Strong membrane expression was observed in HEK293T-CD85A transfected cells, while no staining was observed in the HEK293T cells transfected with the other CD85 family members (CD85B-K). Labelling with the tag anti-V5, anti-MYC and anti-GFP confirmed the efficiency of transfection.

Show image in a new window

Gene inactivation
Not tested
IHC-P Characterisation
Endogenous expression
CD85a (LILRB3/ILT5) antibody IHC-P Characterisation: Endogenous expression
Immunohistochemical staining of the FRAS92B mAb on paraffin-embedded human tissue. CD85A was observed in the paracortical area of the tonsil, the medulla of the thymus and in the red pulp of the spleen. Double immunoenzymatic staining show that CD85A (brown) was not expressed by LCN2-positive granulocytes (red) and CD123-positive plasmacytoid dendritic cells (red).

Show image in a new window

IF Characterisation
Endogenous expression
CD85a (LILRB3/ILT5) antibody IF Characterisation: Endogenous expression
Double immunofluorescence of the FRAS92B mAb (red) with anti-CD68 (green) and Dapi (blue). Note the double labelling of some CD68-positive cells.

Show image in a new window

FC Validation
Over-expression/cross reactivity
Not working
Gene inactivation
Not tested
FC Characterisation
Endogenous expression
Not working
IHC-P Domestic species
Not tested
IHC-P Wild species
Not tested
← Back to CD85a (LILRB3/ILT5) protein