IgA, IgA1, and IgA2 concentrations were determined in 81 defatted human milk samples: colostrum (days 1-5, n = 42), transitional milk (days 6-14, n = 18) and mature milk (days 15-75, n = 21) by immunonephelometry. Correlations were found between total IgA levels and the concentrations of both IgA subclasses (P < 0.0001). The levels of the three molecules decreased over lactation with significant differences (P < 0.05) between colostrum and transitional milk levels and between colostrum and mature milk. Colostral IgA1 and IgA2 mean concentrations dropped respectively from 10.89 +/- 2.12 g/L, and 15.41 +/- 2.10 g/L to 1.83 +/- 0.73 g/L and 3.40 +/- 1.25 g/L in transitional milk reaching finally to 0.36 +/- 0.07 g/L and 0.27 +/- 0.06 g/L in mature milk. IgA2 concentrations were higher than those of IgA1 when the total IgA level was high. The IgA2 levels in colostrum could be an adaptation resistance of IgA to potentially harmful pathogens able to secrete IgA proteases and also a way to regulate colonization of the microflora in the newborn.

Objectives: Microparticle-enhanced nephelometric immunoassays for six human milk proteins (␤-casein, -casein, ␣-lactalbumin, serum albumin, lactoferrin, and lysozyme) and conventional immunonephelometry assays for immunoglobulin A, C3, and C4 complement proteins were developed and characterized. Design and methods: Microparticle-enhanced nephelometric immunoassays are competitive assays based on the nephelometric quantification of the inhibition of microparticle-protein conjugates immunoagglutination by the proteins to be assayed. Results: High precision (CVs ranged from 1% to 14% in within-and between-assays) and recovery (linear recovery in dilution-overloading assay) ensure a reliable determination of the main human milk proteins by single-step homogeneous nephelometric immunoassays, accurate over wide ranges of concentration. These immunoassays were easily applied to a large number of mature human milk samples (between 373 and 503 according to the proteins tested). Conclusions:The immunoassays developed could be applied to the fast determination of human milk protein profile usable for nursery milk bank and fortification.

The mannan binding lectin (MBL) activates the complement system by the lectin pathway after the recognition of some structural motifs (saccharides) present on the surface of microorganisms. MBL has been mostly identified and quantified in human serum by ELISA or microparticle immunonephelometry assays. This article reports the MBL levels as assessed by a microparticle immunonephelometric assay in 76 human milk samples. Immunonephelometry was performed using skim-milk samples diluted 20 times over a calibration range of 0.07-4.82 mg/L. MBL is indeed present in human milk and its concentration decreases significantly during development from colostrum (0.55+/-0.09 mg/L) to transitional (0.18+/-0.02 mg/L) and mature milk (0.17+/-0.02 mg/L). This innate molecule may be involved in the primary defenses of the mammary gland and the neonate, whose immune system is immature. The high levels observed during the first days of lactation support the hypothesis that this molecule plays a key role in limiting the colonization of the newborn gut by pathogens.

The levels of complement fractions C3 and C4 were assayed in human milk in a classic nephelometric assay adapted to this secretion. Concentrations of these molecules were measured in 667 milk samples obtained sequentially from 76 volunteer lactating mothers during the first 12 weeks of lactation. Immunonephelometry was performed using skimmed milk samples diluted 10 times and yielded reproducible (coefficients of variation in within- and between-run precision lower than 9% for C3 and than 14% for C4) and accurate (linear recovery in dilution-overloading assay) data. High concentrations (mean +/- SE) were found for C3 (199.32+/-16.35 mg/L) and C4 (113.42+/-11.16 mg/L) in colostrum samples (n = 159; days 1-5). A significant (P<0.001) and rapid decrease was observed in transitional milk samples (n = 198; days 6-14), containing 57.71+/-5.18 and 72.39+/-4.98 mg/L of C3 and C4, respectively. Stable lower levels were noted in mature milk samples (n = 310; days 15-84) at 30.36+/-1.57 mg/L for C3 (P<0.001) and 53.38+/-3.61 mg/L for C4 (P<0.05). The decrease rate was different for C3 and C4, yielding a reversal of the C3/C4 ratio between colostrum and more mature milk.

Similarly to many immune molecules of human milk, C3 and C4 levels decrease during lactation. We investigated the influence, over the first three weeks of lactation, of both prematurity and parity on the sequential evolution of these levels. Milk C3 and C4 concentrations were measured by immunonephelometry in 494 individual samples collected from 76 lactating mothers. C3 and C4 concentrations were higher in milk from preterm or primiparous mothers. The major differences were observed in milk from days 5-8 and 9-20, likely due to pronounced interindividual variations in levels of days 1-4 milk. Milk from mothers of precocious (33 weeks' gestation or less) preterm newborns presented higher concentrations and a slower decrease of C3 and C4 levels than that from mothers of late (33-37 weeks' gestation) preterm newborns, when compared to term mothers. Finally, the inversion of the C3/C4 ratio occurring over time, previously reported, appeared later in milk from mothers of preterm newborns. The influence of prematurity was even greater in primiparous than in multiparous mothers. Both C3 and C4 levels therefore appear to be influenced in human milk by the parity and prematurity of the delivery. Mothers from preterm newborns seem to provide higher levels of C3 for a longer period post delivery.