The purities and sizes of the fragments were checked by sodium dodecyl sulfate--polyacrylamide gel electrophoresis, which yielded a [F(ab').sub.2] fragment of ~112 kDa and a Fc fragment of 36 kDa (two parts of 18 kDa each).
Available sera from those patients were frozen and thawed and rechecked manually with (a) basic buffer (the screening assay); (b) basic buffer plus 15 mg/L native murine monoclonal MAK33 added; (c) basic buffer plus 15 mg/L heat-treated MAK33; (d) the Fc fragment removed plus basic buffer; (e) the Fc fragment removed plus basic buffer plus 15 mg/L native MAK33; and (f) Fc fragment removed plus basic buffer plus 15 mg/L heat-treated MAK33 (the reference assay).
For type III samples, removal of the Fc fragment normalized CEA concentrations, but not the addition of 15 mg/L heat-treated murine monoclonal MAK33.
Removing the Fc fragment from the capture antibody as the only measure nearly eliminated interference.
A second choice is to reduce interference by removing the Fc fragment (29) or by changing the structure of (32, 33) or chemically modifying (34) the assay antibodies.
Digoxin-specific Fab fragments are produced by cleaving sheep digoxin-specific IgG with papain to form two Fab fragments (50 000 Da each) and one 50 000-Da Fc fragment
The team has engineered antibody Fc fragments
with enhanced sialic acid that target these conserved parts of both HA and NA, binding influenza viruses and thus blocking their interactions with human cells.
IgM antibody by virtue of its large pentameric structure has five Fc fragments
per molecule and will fulfil this requirement.
Nevertheless, use of the intact monoclonal antibodies may provide false positive signals in detection of the tumor cells since the Fc fragments of IgG may bind to the Fc receptors on the surface of normal cells as well.
A number of studies showed that complete IgG has some side effects due to FC fragments. The complete IgG has a long serum half-life resulting in poor contrast during the imaging process; additionally, inappropriate activation of the Fc receptor induces the cross-reactivity, giving rise to associated false positive signals, especially when it is used for FCM [9, 12, 19].
Briefly, Balb/c mice (8-12 weeks of age) were immunized with four intraperitoneal injections of Fc fragments
of IgG myeloma proteins emulsified in Freund's complete adjuvant (Sigma, USA) (first injection) or incomplete adjuvant (Sigma, USA) (other injections) (50 [micro]g, every 2 weeks).
Balb/c mice (8-12 weeks of age) were immunized with four intraperitoneal injections of Fc fragments
of human IgG 1 or IgG2 myeloma proteins emulsified in Freund's complete adjuvant (Sigma, USA) (first injection) or incomplete adjuvant (Sigma) (other injections) (50 [micro]g every 2 weeks).