Separation of ciliary dyskinesia substances found in serum and secreted by cystic fibrosis leukocytes and lymphoid cell lines, using protein A--Sepharose CL-4B.

Cell-free culture medium from PHA-stimulated PBL cultures or long-term lymphoid cell lines 
from CF homozygotes or obligate heterozygotes, as well as serum from 
such individuals, can produce three distinct changes in the ciliary beat 
of rabbit tracheal ciliated epithelium: (1) increased beat frequency, (2) CD, 
and (3) ciliostasis. The ability to produce both 1 and 2 
seems to be characteristic for leukocyte cultures from CF genotypes: 46 
of 49 CF cultures produced both effects, as compared with none 
of 16 cultures from normal controls and 1 of 10 from 
non-CF patient controls with bronchial asthma. CTA was not specific for 
CF PBL cultures, since 13% of normal control and 60% of 
the non-CF patient control cultures produced ciliostasis. Similar findings were obtained 
when serum from the same individuals or culture medium from long-term 
lymphoid cell lines was tested. Using protein A from Staphylococcus aureus 
covalently coupled to Sepharose CL-4B as a reagent, we separated the 
CDAs present in serum and in culture medium from PBL and 
lymphoid cultures. Protein A specifically adsorbs IgG and binds it under 
mild conditions at physiological pH. The CDAs are noncovalently bound to 
IgG at physiological pH and can be co-purified with IgG. The 
IgG-CDA complexes can be readily desorbed from protein A-Sepharose CL-4B at 
pH 3.0, and under these conditions the CDAs are dissociated from 
IgG, retain all their activity, and can be separated by chromatography 
on Bio-Gel P-10 or Sephadex G-200. Three distinct activities were removed 
from serum or PBL cultures which showed both CDA and CTA, 
and two were obtained from samples which showed only CDA. One 
CDA believed to be specific for CF (CF-CDA) was separated from 
CF serum and PBL or lymphoid cell culture medium. This CF-CDA 
was complexed noncovalently to IgG (not subclass IgG3), had a molecular 
weight of 4000 to 6000, was stable as acid pH, was 
found in biological fluids from both homozygotes and heterozygotes for CF, 
was shown not to be C3a (anaphylatoxin), did not require IgG 
for activity, and was positively charged at pH values more acidic 
than 8.0. 