Table 1.

Effect of removal of sialic acid residues and/or removal of polylactosamines on ANCA staining pattern

PatientaSexAge (yr)IIFMPO/PR3
(U/ml)hLAMP-2ANCA
ANCAGEnCNeuraminidasePNGaseFEndo-β-Galactosidase
ANCA-negative/hLAMP-2 autoantibody positive
 AF60++Membrane ++No differenceC+
 BF65NA+Atypical C/
membrane ++No differenceC+
 CF33++Atypical C+No differenceC++
 D (39)M33++Atypical C+No differenceAtypical C++
 EF22++Atypical C++C++Negative
 FF85++CNo difference
 GF70NA+CNo differenceC
 H (70)F17++C++No differenceC++
ANCA-negative/hLAMP-2 autoantibody negative
 IM29NANo differenceNANA
 JF73+No differenceNANA
 KM59+No differenceNo differenceNo difference
ANCA-positive/hLAMP-2 autoantibody negative controls
 LM78PNAMPO 22No differenceNANA
 M (57)F57CNAPR3 20No differenceNo differenceNo difference
 N (51)M58CPR3 50.55No differenceNANA
 O (9)F76CPR3 360No differenceNo differenceNo difference
 PM45CPR3 39.8No differenceNANA
  • All 11 ANCA-negative patients (patients A–K) had negative results in standard, commercially available indirect immunofluorescence assays; however, patient sera with antibodies to hLAMP-2 exhibited binding to GEnC by IIF using either frozen sections of human kidney or immortalized glomerular endothelial cells (n=8, A–H). Using sera from those eight patients for IIF, removal of either N-glycans (n=1, E) or both, removal of sialic acid and polylactosamines (n=7, A–D, F–H) from normal human granulocytes, enhanced fluorescence intensity and revealed either a membrane bound staining (n=2) and/or an (atypical) cANCA pattern (n=7). The remaining three patients did not exhibit hLAMP-2 antibodies by ELISA and treatment of granulocytes did not alter their ANCA negativity by IIF assays (patients I–K). The ANCA staining pattern of five patients with either cANCA/anti-PR3 antibodies (n=4) or pANCA/anti-MPO antibodies (n=1) was not altered by carbohydrate removal (patients L–P), nor did treatment with the enzymes change negative results from sera of healthy controls (n=3). IIF, indirect immunofluorescence; F, female; −, negative; +, positive; membrane ++, membrane accentuated staining; C, cytoplasmic ANCA; NA, not available; M, male; atypical, atypical staining pattern; no difference, no difference in staining pattern to untreated cells; P, perinuclear ANCA.

  • a Numbers in parentheses next to the patients’ assignation (A–P) refer to patients in Supplemental Table 3 published previously by Kain et al. (18).