PMC

Specificity of sELISA. A. The indicated amounts (39 to 625 pg/ml) of synthetic p3-Alc_α35 (diamonds) and p3-Alc_β37 (triangles) peptides were dissolved in buffer A (100 μL per well of a 96-well plate) and assayed with ELISA. B. The indicated amounts (39 to 625 pg/ml) of synthetic p3-Alc_α35 (diamonds) and p3-Alc_α39 (triangles) peptides were dissolved in buffer A (100 μL per well of a 96-well plate) and assayed with ELISA and cross-reactivity was examined. The amino acid sequence information of p3-Alc_α35 (Additional file , Figure S1A) and p3-Alc_β37 are described previously (4, 5).

Levels of p3-Alc_α and Aβ in plasma from clinical populations (Japanese cohort 2). Plasma samples from AD subjects (n = 39) and age-matched nondemented controls (n = 21) were analyzed for levels of p3-Alc_α (A) and Aβ40 (B). Total subjects (All, upper panels), female subjects (Female, middle panels) and male subjects (Male, lower panels) are respectively indicated. Statistical analysis was performed using the Mann-Whitney U-test. **, P = 0.0017 (A, upper). ***, P = 0.0006 (A, middle). ***, P < 0.0001 (B, upper). ***, P < 0.0001 (B, middle). *, P = 0.0263 (B, lower).

Diurnal changes of serum p3-Alc_α levels over a 24 hr period. Sera of young healthy volunteers (A, 6 males; B, 7 females) were examined for p3-Alc_α contents every 8 h within a day. Sampling times (5 p.m., 1 a.m. and 9 a.m.) are indicated. Clinical information about the volunteers and values for p3-Alc_α levels in sera are described in panel C.

Levels of p3-Alc_α and Aβ in plasma from "low Aβ40" and "high Aβ40" populations of AD and FTLD subjects (Japanese cohort 1). Subjects with AD and FTLD were separated into two populations, who showed low (< 340 pg/mL) and high (> 340 pg/ml) Aβ40 levels (cut-off line is shown in the middle panels with dotted line). AD (A) and FTLD (B) are respectively indicated in two populations for p3-Alc_α (left), Aβ40 (middle) and Aβ42 (right). Statistical analysis was performed using the Mann-Whitney U-test. *, P < 0.05; **, P < 0.01; ***, P < 0.001. N.S, not significant.

Correlation between p3-Alc_α and Aβ40 levels in AD and normal subjects (Japanese cohort 2). Correlation of plasma p3-Alc_α and Aβ levels was explored in AD subjects (A) and normal controls (B). Total subjects (All, upper panels), female subjects (Female, middle panels) and male subjects (Male, lower panels) are respectively indicated. Statistical analysis was performed by using the Pearson's correlation coefficient. A. Upper; r = 0.4405 (P = 0.0050), Middle; r = 0.6149 (P = 0.0014), Lower; r = 0.07328 (P = 0.7952). B. Upper; r = 0.6275 (P = 0.0023), Middle; r = 0.6117 (P = 0.0201), Lower; r = 0.7857 (P = 0.0480).

Effect of p3-Alc_α extraction from serum and plasma on the quantification of p3-Alc_α with sELISA. A. Quantification of p3-Alc_α35 peptide in human serum (left) and plasma (right) without extraction. Synthetic p3-Alc_α35 peptide was dissolved in human serum and plasma and the samples in the indicated concentrations, and samples were assayed by ELISA alongside samples in which standard p3-Alc_α35 peptide was dissolved in buffer A (closed diamonds in each respective panel). The p3-Alc_α35 peptide was added to undiluted serum or plasma [closed squares], or to serum or plasma diluted 2-fold [open triangles] or 4-fold [open circles] with buffer A. B. p3-Alc_α35 peptide was added to undiluted serum (left) or plasma (right), and extracted using the procedure described in "Materials and Methods ". Extracted samples (open squares in panel B) were also examined alongside a standard p3-Alc_α35 peptide dissolved in buffer A (closed diamonds). Endogenous p3-Alc_α levels in serum and plasma (no addition of synthetic p3-Alc_α) were subtracted in order to determine the specific curve for quantifying levels of synthetic p3-Alc_α.