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The effects of anti-insulin antibodies and cross-reactivity with human recombinant insulin analogues in the E170 insulin immunometric assay.

Kim S, Yun YM, Hur M, Moon HW, Kim JQ - Korean J Lab Med (2011)

Bottom Line: To investigate the effects of IAs, we performed IA radioimmunoassays, and analyzed the differences between directly measured insulin (direct insulin) and polyethylene glycol (PEG)-treated insulins (free, IA-unbound; total, IA-bound and unbound insulin).In IA-positive patients, E170 free insulin levels measured using the E170 analyzer were significantly lower than the direct insulin levels.Our results suggest that the measurement of free insulin after PEG pre-treatment could be useful for beta cell function assessment in diabetic patients undergoing insulin therapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Laboratory Medicine, Konkuk University School of Medicine, Seoul, Korea.

ABSTRACT

Background: Insulin assays are affected by varying degrees of interference from anti-insulin antibodies (IAs) and by cross-reactivity with recombinant insulin analogues. We evaluated the usefulness of the E170 insulin assay by assessing IA effects and cross-reactivity with 2 analogues.

Methods: Sera were obtained from 59 type 2 diabetes patients receiving continuous subcutaneous insulin infusion and 18 healthy controls. Insulin levels were determined using an E170 analyzer. To investigate the effects of IAs, we performed IA radioimmunoassays, and analyzed the differences between directly measured insulin (direct insulin) and polyethylene glycol (PEG)-treated insulins (free, IA-unbound; total, IA-bound and unbound insulin). We performed in-vitro cross-reactivity tests with insulin aspart and insulin glulisine.

Results: In IA-positive patients, E170 free insulin levels measured using the E170 analyzer were significantly lower than the direct insulin levels. The mean value of the direct/free insulin ratio and IA-bound insulin, which were calculated as the difference between total and free insulin, increased significantly as endogenous IA levels increased. The E170 insulin assay showed low cross-reactivities with both analogues (< 0.7%).

Conclusions: IAs interfered with E170 insulin assay, and the extent of interference correlated with the IA levels, which may be attributable to the increase in IA-bound insulin, and not to an error in the assay. The E170 insulin assay may measure only endogenous insulin since cross-reactivity is low. Our results suggest that the measurement of free insulin after PEG pre-treatment could be useful for beta cell function assessment in diabetic patients undergoing insulin therapy.

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Related in: MedlinePlus

The tendency of E170 direct, total and free insulin levels and fasting glucose level toward the higher anti-insulin antibody level group. The bars are in order of direct, total, and free insulin determined by the E170 insulin assay. The line graph shows the mean level of fasting glucose. Severe insulin resistance appeared in the IA positive DM group 3 (IA>40.0%).*IA(-) DM is the anti-insulin antibody negative group of diabetes patients; †groups 1-3 are groups with anti-insulin antibody positive diabetes patients, which were classified according to IA levels; ‡P values were determined by using Friedman test.Abbreviations: IA, anti-insulin antibody; DM, diabetes mellitus.
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Figure 1: The tendency of E170 direct, total and free insulin levels and fasting glucose level toward the higher anti-insulin antibody level group. The bars are in order of direct, total, and free insulin determined by the E170 insulin assay. The line graph shows the mean level of fasting glucose. Severe insulin resistance appeared in the IA positive DM group 3 (IA>40.0%).*IA(-) DM is the anti-insulin antibody negative group of diabetes patients; †groups 1-3 are groups with anti-insulin antibody positive diabetes patients, which were classified according to IA levels; ‡P values were determined by using Friedman test.Abbreviations: IA, anti-insulin antibody; DM, diabetes mellitus.

Mentions: IA positive diabetes patients were classified into 3 groups according to IA levels. To assess interference from IAs, E170 direct and free insulin levels were compared among IA positive subjects and within each group. E170 free insulin levels were significantly lower than direct insulin levels for IA positive subjects (13.6±15.3 vs. 47.9±67.8, P<0.001 by Wilcoxon test) and this decrease was observed in each of groups (all, P<0.005, Table 2). The mean values of the E170 direct/free insulin ratio, which signified the extent of interference, and of IA-bound insulin levels, which were calculated from the difference between the E170 total and free insulin levels, gradually increased in the higher IA groups, as did the E170 direct, total, and free insulin (Table 2). The differences among the 4 DM groups and total the 5 groups were statistically significant (all, P<0.001 by Kruskal-Wallis H test). The differences in C-peptide and fasting glucose levels among the 5 groups were significant (P=0.031 and 0.003, respectively by one-way ANOVA), but not among the 4 DM groups (P=0.053 and 0.183, respectively). Fasting glucose levels were significantly higher in the IA negative group than in all of the IA positive patients (P=0.046 by Student's t test). The differences in HbA1C and CSII duration were not statistically significant among the 5 groups (P=0.330 and 0.146, respectively), however, CSII duration was significantly shorter in the IA negative group than in all of the IA positive patients (10.2±16.6 vs. 26.1±15.8, P=0.027). Differences among E170 direct, total and free insulin levels were assessed within each of the 4 DM groups (Fig. 1), and were significant only within the IA positive DM groups 2 and 3 (IA negative DM group, P=0.230; IA positive DM group 1, P=0.060; group 2, P=0.023; and group 3, P<0.001, by Friedman test). However, E170 direct insulin levels were not significantly different from E170 total insulin levels (within IA-positive DM group 2, P=0.620; group 3, P=0.092, by Wilcoxon signed rank test; for all subjects, P=0.235, by paired t-test).


The effects of anti-insulin antibodies and cross-reactivity with human recombinant insulin analogues in the E170 insulin immunometric assay.

Kim S, Yun YM, Hur M, Moon HW, Kim JQ - Korean J Lab Med (2011)

The tendency of E170 direct, total and free insulin levels and fasting glucose level toward the higher anti-insulin antibody level group. The bars are in order of direct, total, and free insulin determined by the E170 insulin assay. The line graph shows the mean level of fasting glucose. Severe insulin resistance appeared in the IA positive DM group 3 (IA>40.0%).*IA(-) DM is the anti-insulin antibody negative group of diabetes patients; †groups 1-3 are groups with anti-insulin antibody positive diabetes patients, which were classified according to IA levels; ‡P values were determined by using Friedman test.Abbreviations: IA, anti-insulin antibody; DM, diabetes mellitus.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3111036&req=5

Figure 1: The tendency of E170 direct, total and free insulin levels and fasting glucose level toward the higher anti-insulin antibody level group. The bars are in order of direct, total, and free insulin determined by the E170 insulin assay. The line graph shows the mean level of fasting glucose. Severe insulin resistance appeared in the IA positive DM group 3 (IA>40.0%).*IA(-) DM is the anti-insulin antibody negative group of diabetes patients; †groups 1-3 are groups with anti-insulin antibody positive diabetes patients, which were classified according to IA levels; ‡P values were determined by using Friedman test.Abbreviations: IA, anti-insulin antibody; DM, diabetes mellitus.
Mentions: IA positive diabetes patients were classified into 3 groups according to IA levels. To assess interference from IAs, E170 direct and free insulin levels were compared among IA positive subjects and within each group. E170 free insulin levels were significantly lower than direct insulin levels for IA positive subjects (13.6±15.3 vs. 47.9±67.8, P<0.001 by Wilcoxon test) and this decrease was observed in each of groups (all, P<0.005, Table 2). The mean values of the E170 direct/free insulin ratio, which signified the extent of interference, and of IA-bound insulin levels, which were calculated from the difference between the E170 total and free insulin levels, gradually increased in the higher IA groups, as did the E170 direct, total, and free insulin (Table 2). The differences among the 4 DM groups and total the 5 groups were statistically significant (all, P<0.001 by Kruskal-Wallis H test). The differences in C-peptide and fasting glucose levels among the 5 groups were significant (P=0.031 and 0.003, respectively by one-way ANOVA), but not among the 4 DM groups (P=0.053 and 0.183, respectively). Fasting glucose levels were significantly higher in the IA negative group than in all of the IA positive patients (P=0.046 by Student's t test). The differences in HbA1C and CSII duration were not statistically significant among the 5 groups (P=0.330 and 0.146, respectively), however, CSII duration was significantly shorter in the IA negative group than in all of the IA positive patients (10.2±16.6 vs. 26.1±15.8, P=0.027). Differences among E170 direct, total and free insulin levels were assessed within each of the 4 DM groups (Fig. 1), and were significant only within the IA positive DM groups 2 and 3 (IA negative DM group, P=0.230; IA positive DM group 1, P=0.060; group 2, P=0.023; and group 3, P<0.001, by Friedman test). However, E170 direct insulin levels were not significantly different from E170 total insulin levels (within IA-positive DM group 2, P=0.620; group 3, P=0.092, by Wilcoxon signed rank test; for all subjects, P=0.235, by paired t-test).

Bottom Line: To investigate the effects of IAs, we performed IA radioimmunoassays, and analyzed the differences between directly measured insulin (direct insulin) and polyethylene glycol (PEG)-treated insulins (free, IA-unbound; total, IA-bound and unbound insulin).In IA-positive patients, E170 free insulin levels measured using the E170 analyzer were significantly lower than the direct insulin levels.Our results suggest that the measurement of free insulin after PEG pre-treatment could be useful for beta cell function assessment in diabetic patients undergoing insulin therapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Laboratory Medicine, Konkuk University School of Medicine, Seoul, Korea.

ABSTRACT

Background: Insulin assays are affected by varying degrees of interference from anti-insulin antibodies (IAs) and by cross-reactivity with recombinant insulin analogues. We evaluated the usefulness of the E170 insulin assay by assessing IA effects and cross-reactivity with 2 analogues.

Methods: Sera were obtained from 59 type 2 diabetes patients receiving continuous subcutaneous insulin infusion and 18 healthy controls. Insulin levels were determined using an E170 analyzer. To investigate the effects of IAs, we performed IA radioimmunoassays, and analyzed the differences between directly measured insulin (direct insulin) and polyethylene glycol (PEG)-treated insulins (free, IA-unbound; total, IA-bound and unbound insulin). We performed in-vitro cross-reactivity tests with insulin aspart and insulin glulisine.

Results: In IA-positive patients, E170 free insulin levels measured using the E170 analyzer were significantly lower than the direct insulin levels. The mean value of the direct/free insulin ratio and IA-bound insulin, which were calculated as the difference between total and free insulin, increased significantly as endogenous IA levels increased. The E170 insulin assay showed low cross-reactivities with both analogues (< 0.7%).

Conclusions: IAs interfered with E170 insulin assay, and the extent of interference correlated with the IA levels, which may be attributable to the increase in IA-bound insulin, and not to an error in the assay. The E170 insulin assay may measure only endogenous insulin since cross-reactivity is low. Our results suggest that the measurement of free insulin after PEG pre-treatment could be useful for beta cell function assessment in diabetic patients undergoing insulin therapy.

Show MeSH
Related in: MedlinePlus