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Lactoferrin

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Lactoferrin is a multi-functional iron-binding glycoprotein found in various exocrine secretions, such as milk, saliva, tears, and nasal secretions. It plays a role in iron transport and regulation, as well as in immune function and antimicrobial activity.

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Lab products found in correlation

Neutrophil elastase, by Abcam (2 mentions) Hmgb1, by R&D Systems (2 mentions) Vectastain abc peroxidase kit, by Vector Laboratories (2 mentions) Hsp70, by BioGenex (2 mentions) Ridascreen, by R-Biopharm (1 mentions) Oc sensor diana, by Eiken Chemical (1 mentions) Easysep direct human neutrophil isolation kit, by STEMCELL (1 mentions) Mmp 9, by Abcam (1 mentions) Interleukin 6 (il 6), by R&D Systems (1 mentions) Transferrin, by Abcam (1 mentions) Tnf α, by R&D Systems (1 mentions) Il 33, by R&D Systems (1 mentions) Mpxhcyto 60k, by Merck Group (1 mentions) Synergy ht 96 well plate reader, by Agilent Technologies (1 mentions) Ecl system, by Merck Group (1 mentions) Cytochalasin b, by Merck Group (1 mentions) Formyl methionyl leucyl phenylalanine (fmlp), by Merck Group (1 mentions) Elastase, by Abcam (1 mentions) Myeloperoxidase mpo, by R&D Systems (1 mentions) Mmp 9, by R&D Systems (1 mentions)

Close spelling variants of this product

Lactoferrin [HLF2] human ELISA kit (2 citations) Anti-lactoferrin (2 citations) Ab200015 human lactoferrin simpleStep ELISA Kit (2 citations) Anti-lactoferrin antibody (2 citations)

10 protocols using lactoferrin

1

Inflammatory Biomarkers in Stool and Blood

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Blood samples and stool specimens were collected within 24 hours of admission. Blood inflammatory biomarkers including highly sensitive C-reactive protein (Beckman Coulter, Inc., Brea, CA, USA), erythrocyte sedimentation rate (ESR) (TEST-1; Alifax, Polverara, Italy), and leukocytes from complete blood count (CBC) (Sysmex, Kobe, Japan), and fecal inflammatory biomarkers including calprotectin (Ridascreen®; R-Biopharm, Darmstadt, Germany), lactoferrin (Abcam; Cambridge, UK), MPO (Abcam), and PMN-e (Abcam) were assessed quantitatively. Fecal leukocytes were examined using microscopy. An average number of leukocytes that was ≥1/high power field was considered positive. Fecal occult blood was examined by an immunochemical occult blood test (OC-Sensor Diana; Eiken Chemical Co., Ltd. Tokyo, Japan). A result that was ≥100 ng/mL was considered positive.
Park Y., Son M., Jekarl D.W., Choi H.Y., Kim S.Y, & Lee S. (2019). Clinical Significance of Inflammatory Biomarkers in Acute Pediatric Diarrhea. Pediatric Gastroenterology, Hepatology & Nutrition, 22(4), 369-376.
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2

Neutrophil Activation and Secretome

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Blood neutrophils of healthy donors were isolated using EasySep™ Direct Human Neutrophil Isolation Kit (Stemcell Technologies). Neutrophils were stimulated with bead based immune-complexes and supernatants harvested after 4 hours. Supernatants were diluted 1:5 and release of MPO (Thermo Fisher), MMP9 and Lactoferrin (Abcam) were measured using the Human Elisa Kits according to manufacturer’s instructions. Cytokines were detected in undiluted supernatant using custom-made multiplex cytokine kit (Thermo Fisher).
Bartsch Y.C., Wang C., Zohar T., Fischinger S., Atyeo C., Burke J., Kang J., Edlow A.G., Fasano A., Baden L.R., Nilles E.J., Woolley A.E., Karlson E.W., Hopke A.R., Irimia D., Fischer E.S., Ryan E.T., Charles R., Julg B.D., Lauffenburger D.A., Yonker L.M, & Alter G. (2021). Humoral signatures of protective and pathological SARS-CoV-2 infection in children. Nature medicine, 27(3), 454-462.
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3

Cytokine and Iron Biomarker Profiling

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Supernatants were centrifugated 10 min, 13,000×g, 4 °C. ELISAs were performed according to manufacturer’s instruction: human Hepcidin (R&D Systems, cat#DY8307-05), IL-6 (R&D Systems, cat#DY602), TNFα (R&D Systems, cat#DY2010), IL-8 (R&D Systems, cat#DY208), lactoferrin (abcam, cat#ab108882), and transferrin (abcam, cat#ab108902).
Hazlett H.F., Hampton T.H., Aridgides D.S., Armstrong D.A., Dessaint J.A., Mellinger D.L., Nymon A.B, & Ashare A. (2020). Altered iron metabolism in cystic fibrosis macrophages: the impact of CFTR modulators and implications for Pseudomonas aeruginosa survival. Scientific Reports, 10, 10935.
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4

Quantification of Inflammatory Markers

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IL-1β, IL-6, IL-7, IL-8 (CXCL8), IL-10, IL-12p70, tumor necrosis factor alpha (TNFα), CXCL1 (growth-related oncogene), CXCL10 (human interferon-inducible protein 10), CCL2 (monocyte chemoattractant protein-1), CCL3 (macrophage inflammatory protein-1α), CCL4 (macrophage inflammatory protein-1β), and CCL22 (macrophage-derived chemokine) were quantified with bead-based multiplex assays (HSCYTO-60SK and MPXHCYTO-60K, Millipore, Billerica, MA). IL-33 (R&D Systems, Minneapolis, MN), transforming growth factor beta-1 (TGF-β1) (Promega, Madison, WI), neutrophil elastase (Abcam, Cambridge, MA), myeloperoxidase (R&D Systems) and lactoferrin (Abcam) ELISA were performed according to manufacturer instructions. Concentrations were determined with a BioTek (Winooski, VT) Synergy HT 96-well plate reader using manufacture recommended wavelengths and standard curve fit.
Grunwell J.R., Stephenson S.T., Tirouvanziam R., Brown L.A., Brown M.R, & Fitzpatrick A.M. (2018). Children with neutrophil-predominant severe asthma have pro-inflammatory neutrophils with enhanced survival and impaired clearance. The journal of allergy and clinical immunology. In practice, 7(2), 516-525.e6.
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5

Neutrophil Degranulation Assay

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Neutrophils (5 × 106/mL) were pre-incubated for 10 min with APPA (100 µg/mL), before priming with GM-CSF (5 ng/mL) for 30 min and then stimulated to degranulate with cytochalasin B (5 µg/mL) plus fMLP (1 µM, both from Sigma) for 15 min. Cells were pelleted gently, washed and analysed by flow cytometry; while, supernatants were removed for SDS-PAGE after adding concentrated Laemmli protein sample buffer. After electrophoresis, proteins were transferred to PVDF membranes and probed with antibodies to myeloperoxidase (MPO) (R&D Systems), lactoferrin (Abcam), MMP9 (R&D Systems) and elastase (Abcam). Secondary antibodies were anti-rabbit IgG (GE Healthcare) and anti-mouse IgG (Sigma) HRP-linked antibodies (1:10,000). Bound antibodies were detected using the ECL system (Merck) and film (Amersham).
Cross A.L., Hawkes J., Wright H.L., Moots R.J, & Edwards S.W. (2020). APPA (apocynin and paeonol) modulates pathological aspects of human neutrophil function, without supressing antimicrobial ability, and inhibits TNFα expression and signalling. Inflammopharmacology, 28(5), 1223-1235.
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6

Salivary Biomarkers Quantification

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Commercial ELISA kits were used to determine the salivary concentrations of immunoglobulin A (IgA) (Salimetrics, Carlsbad, CA, USA) and lactoferrin (Abcam, Cambridge, MA, USA). The salivary lipopolysaccharide (LPS) concentration was determined using a Limulus Amoebocyte Lysate Assay Kit (Associates of Cape Cod, East Falmouth, MA, USA).
Uchida F., Oh S., Shida T., Suzuki H., Yamagata K., Mizokami Y., Bukawa H., Tanaka K, & Shoda J. (2021). Effects of Exercise on the Oral Microbiota and Saliva of Patients with Non-Alcoholic Fatty Liver Disease. International Journal of Environmental Research and Public Health, 18(7), 3470.
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7

Immunohistochemistry of HMGB1, RAGE, and HSP70

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Sections from formalin fixed tissues in paraffin blocks were deparaffinised and rehydrated before microwave-assisted antigen retrieval in citric acid buffer at pH 6.0. Endogenous peroxidase activity was blocked with CH3OH/H2O2 treatment and were blocked with 2% BSA in PBS. Sections were incubated overnight at 4°C with the primary antibody diluted in 2% serum in PBS(7 (link)). We used the following primary antibodies: Lactoferrin (Abcam, Cambridge, MA, dilution 1:50), HMGB1 (R&D Systems, Minneapolis, MN, dilution 1:50), RAGE (AbD Serotec, Oxford, UK, dilution 1:1000), HSP70 (Biogenex, San Ramon, CA, dilution 1:50). Sections were then washed and incubated with the appropriate species-specific secondary antibody diluted 1:200 in 2% serum for 2 hours at room temperature. After further washing, antigen:antibody complexes were visualized using a Vectastain ABC peroxidase kit (Vector Laboratories Inc., Burlingame, CA). Antigen detection was enhanced with nickel-diaminobenzidine, followed by incubation with TRIS-cobalt. Slides were counterstained with Nuclear Fast Red for photomicroscopy.
Schmidt A.F., Kannan P.S., Kemp M.W., Kramer B.W., Newnham J.P., Jobe A.H, & Kallapur S.G. (2014). Intraamniotic LPS modulates expression of antimicrobial peptides in the fetal sheep lung. Pediatric research, 76(5), 441-447.
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8

Immunostaining of Kidney Allograft Biopsies

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Nine representative cases of kidney allograft biopsies of PVAN (n=3), TCMR (n=3) and STA (n=3) were stained for 2 over-expressed transcripts in PVAN kidney biopsies. To determine the extent of renal damage and classify them into the 3 different phenotypes (PVAN, TCMR and STA), all renal biopsies were analyzed by 2 blinded pathologists. The biopsies were probed with rabbit polyclonal to Lactoferrin (Abcam, Cambridge, MA), rabbit polyclonal to IFN-inducible transmembrane 1 (IFITM-1) (Abcam, Cambridge, MA) and SV40 (Becton Dickinson, Madrid, Spain). Immune staining in formalin-fixed, paraffin-embedded tissues was performed as described previously 18 (link)-20 (link). As positive controls for LTF, IFITM1 and SV40 human tonsil, liver carcinoma and kidney allograft paraffin-embedded tissues were used as positive controls, respectively. To quantify LTF and IFITM1 expression a semi quantitative score from 0 to 3 in the different compartments of the kidney (glomeruli, vessels, tubuli and interstitium) was used.
Sigdel T.K., Bestard O., Salomonis N., Hsieh S.C., Torras J., Naesens M., Tran T.Q., Roedder S, & Sarwal M.M. (2016). Intragraft Antiviral-specific Gene Expression as a Distinctive Transcriptional Signature for Studies in Polyomavirus Associated Nephropathy. Transplantation, 100(10), 2062-2070.
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9

Immunohistochemistry of HMGB1, RAGE, and HSP70

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Sections from formalin fixed tissues in paraffin blocks were deparaffinised and rehydrated before microwave-assisted antigen retrieval in citric acid buffer at pH 6.0. Endogenous peroxidase activity was blocked with CH3OH/H2O2 treatment and were blocked with 2% BSA in PBS. Sections were incubated overnight at 4°C with the primary antibody diluted in 2% serum in PBS(7 (link)). We used the following primary antibodies: Lactoferrin (Abcam, Cambridge, MA, dilution 1:50), HMGB1 (R&D Systems, Minneapolis, MN, dilution 1:50), RAGE (AbD Serotec, Oxford, UK, dilution 1:1000), HSP70 (Biogenex, San Ramon, CA, dilution 1:50). Sections were then washed and incubated with the appropriate species-specific secondary antibody diluted 1:200 in 2% serum for 2 hours at room temperature. After further washing, antigen:antibody complexes were visualized using a Vectastain ABC peroxidase kit (Vector Laboratories Inc., Burlingame, CA). Antigen detection was enhanced with nickel-diaminobenzidine, followed by incubation with TRIS-cobalt. Slides were counterstained with Nuclear Fast Red for photomicroscopy.
Schmidt A.F., Kannan P.S., Kemp M.W., Kramer B.W., Newnham J.P., Jobe A.H, & Kallapur S.G. (2014). Intraamniotic LPS modulates expression of antimicrobial peptides in the fetal sheep lung. Pediatric research, 76(5), 441-447.
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10

Comprehensive Analysis of Neutrophil Biology

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Antibodies specific for Ly6G, F4/80, CD35, and rabbit immunoglobulin G (IgG) were from BioLegend; CD11b was from Tonbo; CD63 was from BD Biosciences; and MPO, H3Cit, lactoferrin, PR3, neutrophil elastase, Fas, dsDNA, and mouse IgG alkaline phosphatase were from Abcam. Streptavidin Alexa Fluor 488 was from BioLegend. Live/dead stain and DHR123, CellTracker Orange, CellTrace Violet, CellMask Deep Red, and BacLight Green Bacterial Stain were from Invitrogen; Helix NP Blue was from BioLegend. Murine Fc-blocking antibody (2.4G2) was from Tonbo. PMA and DNase I were ordered from Sigma-Aldrich. Cytochalasin D was purchased from Cayman Chemical. Paraformaldehyde was from Electron Microscopy Sciences. The neutrophil elastase activity kit was purchased from Abcam. Histopaque 1119 and Histopaque 1077 were from Sigma-Aldrich; Lympholyte was from Cedarlane Laboratories. Dulbecco’s modified Eagle’s medium (DMEM) and phosphate-buffered saline (PBS) that were from Gibco and fetal bovine serum (FBS) from Atlanta Biologicals were used for all tissue culture.
Monteith A.J., Miller J.M., Maxwell C.N., Chazin W.J, & Skaar E.P. (2021). Neutrophil extracellular traps enhance macrophage killing of bacterial pathogens. Science Advances, 7(37), eabj2101.
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