age: 44 gender: female platelet count: 222 hematocrit: 36 acute mat titer: 0 convalescent mat titer: 200 elisa: negative pcr: negative lab confirmation: positive mat result: positive number of days of symptoms: 3 respiratory failure: NA icu admission: NA oliguric renal failure: NA
Extracted molecule
protein
Extraction protocol
Human sera samples were diluted 1/100 in Protein Array Blocking Buffer containing E. coli lysate 10 mg/mL (McLab) at a final concentration of 10% v/v and incubated for 30 min at room temperature under constant mixing to remove background reactivity to E. coli proteins in the IVTT reactions. E. coli protein-antibody complexes were removed from the sample dilution mix via centrifugation prior to addition to the microarray
Label
P3
Label protocol
Biotinylated anti-human IgG, diluted 1/2000 in Blocking Buffer or anti-human IgM, diluted 1/400 in Blocking Buffer, was added to the arrays for one-hour incubation at room temperature. Bound antibodies were detected by one-hour incubation with streptavidin-conjugated SureLight P3.
Hybridization protocol
Slides were washed 3 times with TTBS after each incubation
Scan protocol
Slides were scanned in a Perkin Elmer ScanArray confocal laser and intensities were quantified using QuantArray package
Description
-
Data processing
Array signal intensity was quantified using QuantArray software. Spots intensity raw data were obtained as the mean pixel signal intensity with automatic correction forspot-specific background. Data was normalized by dividing the raw signal for each IVTT protein spot by the median of the sample-specific IVTT control spots (fold-overcontrol [FOC]) and then taking the base-2 logarithm of the ratio (log2 FOC).
Antibody profile in patients with mild and severe leptospirosis
Data table header descriptions
ID_REF
VALUE
Conceptually, a normalized signal of 0.0 is equal to control spot signal, and anormalized signal of 1.0 is 2-fold higher than control spot signal. When evaluating a protein spot as reactive or non-reactive, normalized signals >1.0 were considered reactive. These designations were used to evaluate responsefrequency and to identify a subset of sero-reactive proteins for further analysis. A given protein on the array was considered sero-reactive if it was reactive in at least 60% of the samples in one or more of the following groups: severe disease, acute sample (n=30); severe disease, convalescent sample (n=30); mild disease, acute sample (n=30); mild disease, convalescent sample (n=30); endemic controls (n=30); naïve controls (n=30).
