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Page 6 of 19 van Beek et al. Microbiome Res Rep 2025;4:13 https://dx.doi.org/10.20517/mrr.2024.45

parameter logistic curve was used for the calculation of the results.

IAP
IAP activity was quantified with QUANTI-Blue solution (rep-qbs, InvivoGen) according to the
manufacturer’s instructions. A standard test was performed to identify the correct standard concentrations.
Secreted embryonic alkaline phosphatase (SEAP) protein (rec-hseap, Invivogen) was used as the standard.
A linear standard curve was used for the calculation of the results.

BPI
BPI was quantified by sandwich ELISA using BPI, Human, ELISA kit (HK314, Hycult Biotech) according to
the manufacturer’s instructions, except samples were diluted 1:2 in dilution buffer. A four-parameter
logistic curve was used for the calculation of the results.

HD5
Alpha defensin five was quantified with sandwich ELISA using HD5 (Paneth Cell Specific) ELISA Kit (E-
EL-H1798, Elabscience) according to the manufacturer’s instructions. Samples were diluted 1:2 in reference
standard and sample diluent. A four-parameter logistic curve was used for the calculation of the results.

ECP
ECP was quantified with sandwich ELISA using Human RNASE3/ECP (Ribonuclease A3/Eosinophil
Cationic Protein) ELISA Kit (E-EL-H1379, Elabscience) according to the manufacturer’s instructions.
Samples were diluted 1:2 in reference standard and sample diluent. A four-parameter logistic curve was
used for the calculation of the results.

Muc2
Muc2 was quantified by sandwich ELISA using Human Muc2 ELISA Kit (E-EL-H0632, Elabscience)
according to the manufacturer’s protocol. Samples were diluted 1:4 in reference standard and sample
diluent. A four-parameter logistic curve was used for the calculation of the results.

IgA
IgA was quantified by sandwich ELISA using Human IgA Matched Antibody Pair Kit (Abcam, ab219536)
according to the manufacturer’s general protocol for matched antibody pair kits. A linear standard curve
was used for the calculation of the results.

LTF
LTF was quantified with sandwich ELISA using Human Lactotransferrin / LTF ELISA Pair Set (SEK11096,
SinoBiological) according to the manufacturer’s instructions. A linear standard curve was used for the
calculation of the results.

LCN2
LCN2 was quantified with sandwich ELISA using Human Lipocalin-2/NGAL Matched ELISA Antibody
Pair Set (SEK10222, SinoBiological) according to the manufacturer’s instructions, using 100 μL of STOP
solution per sample. A linear standard curve was used for the calculation of the results.

Cal
Cal was quantified by sandwich ELISA using Human Cal (S100A8 + S100A9) Antibody Pair - BSA and
Azide free (Abcam, ab309558) and Recombinant Human Cal (S100A8 + S100A9) protein (Abcam,