All available sera from residual specimens from NHANES 2013-2014 participants were tested for Toxocara spp. by a Luminex assay using recombinant rTc-CTL-1 antigen that detects IgG antibodies against Toxocara spp. All results in this data release are reported as positive or negative.
All specimens from males and females aged 6+ from 2013-2014 leftover specimens from laboratories that tested them and sent the residual serum back to the NCHS repository.
Toxocara Assay Procedure in Luminex platform:
MagPlex Immunoassay
Fifty μL of working microsphere mixture (50 beads/μL in PBS/0.3% Tween-20/5% non-fat dry skim milk) and 50 μL of diluted sera (1:100 dilution in PBS/0.3% Tween-20/5% non-fat dry skim milk) were added into each well of Costar 96-well black, round-bottom plate (Fisher Scientific, Cat.# 3792). After 30 minutes incubation at room temperature with shaking at speed 6 (~800 rpm), the beads were washed using Biotek Magnetic Washer ELx50 (2 minutes magnetic separation, and then 2 cycles of dispensing 100 µL of PBS/Tween-20 0.3%, soaks for 40 seconds before aspiration). The complex of antibody and coupled beads was detected using 50 μL of biotinylated mouse anti-human IgG (clone H2, affinity purified, Southern Biotech, Birmingham, AL, Cat. Number 9042-08) diluted 1:200 in PBS-1% BSA, 0.05% NaN3. After 30 minutes incubation, the beads were washed as before. As detector, 50 μL/well of diluted Streptavidin, R-phycoerythrin conjugate (Invitrogen, Cat. # S866) (1:250 dilution in PBS-1% BSA, 0.05% NaN3) was added to the well and incubated for another 30 minutes. After washing, the beads were resuspended using 100 μL/well of PBS-1% BSA, 0.05% NaN3. The mean fluorescence intensity from each well was determined by using BioPlex manager software, version 6.02 (BioRad) in Luminex 100 platform. The mean fluorescence intensity – signal intensity of the blank well is used for further analysis.
Quality control and assay validation information:
For each run, we ran three controls: Toxocara 100, Toxocara 50, and Toxocara 0 (negative). These 3 controls should be within the mean ± 2 standard deviations of the established value. If the controls of the plate have a value outside of the acceptance range, the whole plate was retested. The assay itself has been validated against reference serum sets and it has a sensitivity of 90% and a specificity of 99%.
N/A
The test was positive if the antibody response to rTc-CTL-1 was greater than the cut-off point value (23.1 mean fluorescence intensity [MFI]) and negative test if the response was ≤ 23.1 MFI.
Code or Value | Value Description | Count | Cumulative | Skip to Item |
---|---|---|---|---|
-9 to 20830.5 | Range of Values | 6578 | 6578 | |
. | Missing | 0 | 6578 |
Code or Value | Value Description | Count | Cumulative | Skip to Item |
---|---|---|---|---|
1 | Positive | 371 | 371 | |
2 | Negative | 6207 | 6578 | |
. | Missing | 0 | 6578 |