Ann Lab Med.  2015 May;35(3):321-328. 10.3343/alm.2015.35.3.321.

Interlaboratory Comparison of the Results of Lifecodes LSA Class I and Class II Single Antigen Kits for Human Leukocyte Antigen Antibody Detection

• Oh EJ
• Park H
• Park KU
• Kang ES
• Kim HS
• Song EY

• KMID: 2363204
• DOI: http://doi.org/10.3343/alm.2015.35.3.321

Abstract

BACKGROUND
Although single antigen bead assays (SAB) are approved qualitative tests, the median fluorescence intensity (MFI) values obtained from SAB are frequently used in combination with quantitative significances for diagnostic purposes. To gauge the reproducibility of SAB results, we assessed the interlaboratory variability of MFI values using identical kits with reagents from the same lot and the manufacturer's protocol.
METHODS
Six serum samples containing HLA-specific antibodies were analyzed at five laboratories by using Lifecodes LSA Class I and Class II SAB kits (Immucor, USA) from the same lot, according to the manufacturer's protocol. We analyzed the concordance of qualitative results according to distinct MFI cutoffs (1,000, 3,000, 5,000, and 10,000), and the correlation of quantitative MFI values obtained by the participating laboratories. The CV for MFI values were analyzed and grouped by mean MFI values from the five laboratories (<1,000; 1,000-2,999; 3,000-4,999; 5,000-9,999; and > or =10,000).
RESULTS
The categorical results obtained from the five laboratories exhibited concordance rates of 96.0% and 97.2% for detection of HLA class I and class II antibodies, respectively. The Pearson correlation coefficients for MFI values of class I and class II antibodies were between 0.947-0.991 and 0.992-0.997, respectively. The median CVs for the MFI values among five laboratories in the lower MFI range (<1,000) were significantly higher than those for the other MFI ranges (all P<0.01).
CONCLUSIONS
Analysis of SAB performed in five laboratories using identical protocols and reagents from the same lot resulted in high levels of concordance and strong correlation of results.

Keyword

Antibody specificity; Correlation; HLA antigens; Histocompatibility testing; Laboratories; Concordance

MeSH Terms

Analysis of Variance
HLA Antigens/immunology
Histocompatibility Testing
Humans
Isoantibodies/*blood
Laboratories
Reagent Kits, Diagnostic
Reproducibility of Results
HLA Antigens
Isoantibodies
Reagent Kits, Diagnostic

Figure

• 

  Fig. 1 Concordance of single antigen bead assay (SAB) results among the five participating laboratories. SAB analysis was performed by using six serum samples and four different median fluorescence intensity (MFI) cutoffs. The X-axis shows the different MFI cutoffs, and the Y-axis shows the frequency of concordant results. (A) Class I; (B) Class II beads.

• 

  Fig. 2 Box plots (minimum, first quartile, median value, third quartile, and maximum value) of median fluorescence intensity (MFI) values from single antigen bead assays (SAB) that yielded inconsistent results among the five participating laboratories. Results are grouped by the different MFI cutoff values (1,000, 3,000, 5,000, and 10,000); upper panel contains data for HLA class I antibody detection; lower panel contains data for HLA class II antibody detection.*P<0.05; **P<0.01 by Mann-Whitney test.

• 

  Fig. 3 Box plots (minimum, first quartile, median value, third quartile, and maximum value) of CV (%) for median fluorescence intensity (MFI) values separated by distinct MFI ranges and grouped by the mean MFI values from the five participating laboratories (<1,000; 1,000-2,999; 3,000-4,999; 5,000-9,999; and ≥10,000). Asterisk means outlier greater than 3 times the interquartile range. Open circle means outlier greater than 1.5 times the interquartile range. (A) contains data for HLA class I antibody detection; (B) contains data for HLA class II antibody detection.

• 

  Fig. 4 Box plots (minimum, first quartile, median value, third quartile, and maximum value) of CV (%) for median fluorescence intensity (MFI) values, according to HLA antigens (HLA-A, B, C, DR, DQ, and DP). Asterisk means outlier greater than 3 times the interquartile range. Open circle means outlier greater than 1.5 times the interquartile range.

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