Published work relying on immunoGlobe services (selection):

  • Wild et al. (2011) Science 333:228-33.
  • Günther et al. (2009) J. Mol. Med. 87:31-41.
  • Pagenstecher et al. (2009) Hum. Mol. Genetics 18: 911–918.
  • Fritsche et al. (2008) Nature Genetics 40: 892-896.
  • Agerer et al. (2003) J. Biol. Chem. 278,: 42524–42531.
  • Butt et al. (2003) J. Biol. Chem. 278,: 15601-15607.

Different ways to "purify" an antibody

Strictly speaking, not all "purified" or "affinity purified" antibodies on the market are actually affinity purified. Equivalently, not all methods of antibody purifications are reasonable for all purposes. Here we explain the differences and we compare the different grades of antibody purity:

Designation Method of Purification Purity PurposeReduces background in WB, IF/IHC ?
crude serum
-

total serum

Percentage of specific antibodies

  • (as a fraction of total serum protein): 0.02%-0.1%-(1%)(depending on titre)
  • (as a fraction of total immunoglobulins): 0.1%-0.5%-(5%)(depending on titre)

other serum proteins (albumin and others) prevent efficient plate coating and labeling

-

'purified' /total IgG
e.g. ammonium sulfate precipitation

total immunoglobulins (irrespective of specificity), often contaminated by high molecular weight proteins and other contaminants

Percentage of specific antibodies: 0.1%-0.5%-(5%)(depending on titre)

conditionally useful: for plate coating and direct labeling

no

'purified' /total IgG ammoinium sulfate ppt. + gel filtration

total immunoglobulins (irrespective of specificity), less impurities than without gel filtration

Percentage of specific antibodies: 0.1%-0.5%-(5%)(depending on titre)

conditionally useful: for plate coating and direct labeling no
purified, ('affinity purified') protein A / protein G

total IgG (irrespective of specificity) extracted from serum or cell culture supernatant based on the Fc (antibody constant part) binding affinity of protein A or G. Therefore these antibodies sometimes misleadingly are called "affinity purified"

Percentage of specific antibodies:

  • polyclonal (pABs)

    0.1%-0.5%-(5%)(depending on titre)
  • monoclonal (mABs):

    ≤ 100% (provided that measures are taken to to prevent co-purification of bovine IgG derived from FCS)

a) pABs:

conditionally useful: for plate coating and direct labeling

 

 

 

b) mABs:

perfect for

any application

(labelling, coating, IF/IHC, WB, IP ...)

no

affinity purified antigen matrix

highly purified antibodies

Percentage of specific antibodies:

≤ 100%

perfect for any application

(labelling, coating, IF/IHC, WB, IP ...)

yes