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Emax immunoassay system

Manufactured by Promega
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The Emax ImmunoAssay System is a high-performance microplate reader designed for various immunological assays. It offers precise absorbance measurements, making it suitable for applications such as ELISA, cell-based assays, and other immunoassay techniques.

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Lab products found in correlation

Bca protein assay kit, by Thermo Fisher Scientific (2 mentions) Protease inhibitor cocktail, by Thermo Fisher Scientific (2 mentions) Aprotinin, by Santa Cruz Biotechnology (2 mentions) Leupeptin, by Santa Cruz Biotechnology (2 mentions) N per neuronal protein extraction reagent, by Thermo Fisher Scientific (2 mentions) Synergy mx, by Agilent Technologies (2 mentions) Phenyl methyl sulfonyl fluoride (pmsf), by Merck Group (1 mentions) Dopamine elisa kit, by Abnova (1 mentions) Anti hb egf antibody, by R&D Systems (1 mentions) Elisa kit, by USCN (1 mentions) Liaison autoanalyzer, by DiaSorin (1 mentions) Quantikine sandwich elisa kit, by R&D Systems (1 mentions) Human aβ40 elisa kit, by Thermo Fisher Scientific (1 mentions) Human aβ42 elisa kit, by Thermo Fisher Scientific (1 mentions) Protease inhibitor cocktail, by Merck Group (1 mentions) Benchmark microplate reader, by Bio-Rad (1 mentions) Bdnf emax immunoassay system, by Promega (1 mentions) P0044, by Merck Group (1 mentions) P5726, by Merck Group (1 mentions) P8340, by Merck Group (1 mentions)

31 protocols using emax immunoassay system

1

Brain BDNF Quantification by ELISA

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To determine BDNF content in the cortex and hippocampus, the ELISA kit Emax® ImmunoAssay System was used (ref G7611, Promega, USA). The assay was performed according to the manufacturer’s instructions.
Morzelle M.C., Salgado J.M., Telles M., Mourelle D., Bachiega P., Buck H.S, & Viel T.A. (2016). Neuroprotective Effects of Pomegranate Peel Extract after Chronic Infusion with Amyloid-β Peptide in Mice. PLoS ONE, 11(11), e0166123.
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2

Bdnf Quantification in Hippocampus

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Brain tissues were obtained as described for Western blot and stored at −80 °C. Prior to analysis, samples were thawed and then weighed. Lysis buffer (100 mM PIPES pH 7, 500 mM NaCl, 0.2% Triton X-100, 2% BSA, 2 mM EDTA, 200 μM PMSF, 1 × protease, phosphatase-1, and phosphatase-2 inhibitor cocktails from Sigma) was then pipetted into each tube (100 μl per mg of tissue for each left hippocampus). Samples were homogenized, sonicated and centrifuged for 30 min at 16,000 × g at 4 °C. Supernatants were then removed and frozen at −80 °C until analysis. The concentration of Bdnf was determined using the E-Max ImmunoAssay system (Promega) according to the manufacturer’s instructions.
Podda M.V., Cocco S., Mastrodonato A., Fusco S., Leone L., Barbati S.A., Colussi C., Ripoli C, & Grassi C. (2016). Anodal transcranial direct current stimulation boosts synaptic plasticity and memory in mice via epigenetic regulation of Bdnf expression. Scientific Reports, 6, 22180.
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3

Quantification of Urinary Biomarkers

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Women provided a first morning urine sample within one week of completing the study questionnaires. Voided urine was received in cold mailers and ice packs, and transferred to the laboratory for analysis. The samples were centrifuged at 3,000 rpm for 10 minutes at 4°C. The supernatant was separated into 1.5 ml aliquots, and preserved in −80°C freezer.
Urinary NGF and BDNF concentrations were determined using the Emax ImmunoAssay System (Promega, Madison, WI, USA), a precise and highly sensitive ELISA kit, which had a minimum sensitivity of 7.8 pg/ml. Urinary Osteopontin (Opn) levels were determined using Human Osteopontin EIA kit (Enzo Life Sciences, Farmingdale, NY) which had a minimum sensitivity of 0.110 ng/ml. Urinary VEGF levels were measured using Human VEGF EIA kit (Thermo Scientific, Waltham, MA) with a minimum sensitivity of 5pg/ml. All assays were performed according to the manufacturer’s instructions. The amounts of urinary biomarkers in each urine sample were calculated based on a standard curve. All samples were run in duplicate, and the values were averaged. Similar to prior studies, total urinary NGF, BDNF, VEGF and Opn levels were normalized to urinary creatinine (mg/dl) concentration (NGF/Cr, BDNF/Cr, VEGF/Cr and Opn/Cr levels) for each individual.22 (link)
Soriano A., Andy U., Hassani D., Whitmore K., Harvie H., Malykhina A.P, & Arya L. (2021). Relationship of bladder pain with clinical and urinary markers of neuroinflammation in women with urinary urgency without urinary incontinence. Female pelvic medicine & reconstructive surgery, 27(2), e418-e422.
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4

Measuring Neurotrophic Factors and Dopamine in Mouse Brains

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The levels of BDNF, GDNF, and NGF were measured as previously described [31 (link)]. The PBS-perfused cerebral cortex was excised from each mouse and homogenized. Protein levels were measured using a two-site sandwich ELISA (Emax Immunoassay System, Promega, Madison, WI, USA). Dopamine levels in the mouse brains were measured using a Dopamine ELISA kit (KA1887, Abnova Corp, Taiwan), according to the manufacturer’s instruction. The protein concentration in each sample was measured using a BCA protein assay kit (Thermo Fisher Scientific).
Nakamura T.Y., Nakao S., Nakajo Y., Takahashi J.C., Wakabayashi S, & Yanamoto H. (2017). Possible Signaling Pathways Mediating Neuronal Calcium Sensor-1-Dependent Spatial Learning and Memory in Mice. PLoS ONE, 12(1), e0170829.
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5

Measuring Urinary NGF and HB-EGF Levels

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Urinary NGF levels were measured using the Emax® Immuno Assay System (Promega, Madison, WI, USA) with an ELISA kit (USCN Life Science, Wuhan, Hubei, China). Assays were performed according to the manufacturer's instructions, as previously reported.5 (link) For the HB-EGF ELISA, each well of a 96-well Immulon II plate (Dynatech Laboratories, Chantilly, VA, USA) was coated with 200 µL of urine at 4℃, overnight, as previously reported.14 (link) The plates were blocked with 5% fetal bovine serum, and anti-HB-EGF antibody (1 µg/mL) (R&D Systems, Minneapolis, MN, USA) was added in addition to biotinylated anti-goat IgG/avidin D-horseradish peroxidase. The total urinary NGF and HB-EGF levels were further normalized to the concentration of urinary creatinine (NGF/Cr and HB-EGF/Cr levels).
Kim S.R., Moon Y.J., Kim S.K, & Bai S.W. (2014). NGF and HB-EGF: Potential Biomarkers that Reflect the Effects of Fesoterodine in Patients with Overactive Bladder Syndrome. Yonsei Medical Journal, 56(1), 204-211.
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6

Measuring Serum BDNF and IGF-I Levels

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Brain-derived neurotrophic factor (BDNF) protein levels were measured in serum samples using the Emax Immuno Assay system from Promega (Madison, WI, USA). IGF-I (nmol/l) was assayed centrally by chemiluminescence immunoassay of EDTA plasma on the Liaison autoanalyzer (DiaSorin, S.p.A., Italy). Both measures have previously been found to be associated with depression with melancholic features in this sample (Patas et al., 2014 ).
Beijers L., Wardenaar K.J., Bosker F.J., Lamers F., van Grootheest G., de Boer M.K., Penninx B.W, & Schoevers R.A. (2018). Biomarker-based subtyping of depression and anxiety disorders using Latent Class Analysis. A NESDA study. Psychological Medicine, 49(4), 617-627.
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7

Neurochemical Quantification in Hippocampus

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The hippocampus was homogenized in a lysis buffer containing 0.05 M Tris-HCl pH 7.5, 0.15 M NaCl, 1% NP-40, 1 mM Na-orthovanadate, 0.001% sodium fluoride, 1% protease inhibitor cocktail (Sigma). BDNF, NT3, and NGF were assayed using the Emax® immunoassay system (Promega) and FGF2 and IGF1 were assayed using the Quantikine® sandwich ELISA kit (R&D Systems) according to manufacturer’s instructions and as described in our previous publications [38] (link)–[41] (link). Aβ40 and Aβ42 levels were assayed using Aβ40 Human ELISA Kit and Aβ42 Human ELISA Kit according to the manufacturer’s instructions (Invitrogen).
Mellott T.J., Pender S.M., Burke R.M., Langley E.A, & Blusztajn J.K. (2014). IGF2 Ameliorates Amyloidosis, Increases Cholinergic Marker Expression and Raises BMP9 and Neurotrophin Levels in the Hippocampus of the APPswePS1dE9 Alzheimer’s Disease Model Mice. PLoS ONE, 9(4), e94287.
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8

Quantifying BDNF Levels in Hippocampal Samples

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Aliquots of crude bilateral hippocampal homogenates for Western blot assay were stored at -80°C. The Western blot was carried out as described earlier (Radak et al, 2006 (link)). Samples were incubated in block solution supplemented with primary antibody against BDNF, diluted 1:5000 (polyclonal rabbit anti-BDNF Santa Cruz SC-546) overnight at 4°C with constant agitation. To normalize the expression of BDNF, the blots were re-incubated with a mouse anti-GAPDH antibody. Antibody anti β-actin (monoclonal mouse Cat# sc-8035, Santa Cruz) was added to the incubation solution for endogenous control. The concentration of BDNF was determined from the hippocampal section of the brain, using the E-Max ImmunoAssay System (Promega, Madison, WI) as described previously (Radak et al, 2013b (link)).
Torma F., Bori Z., Koltai E., Felszeghy K., Vacz G., Koch L., Britton S., Boldogh I, & Radak Z. (2014). Eating habits modulate short term memory and epigenetical regulation of brain derived neurotrophic factor in hippocampus of low-and high running capacity rats. Brain research bulletin, 0, 54-60.
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9

Serum BDNF Concentration Measurement Protocol

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As described in more detail elsewhere,34 (link) serum was separated immediately after blood draw and stored (at −85 °C) until assay. Serum BDNF concentrations were determined using the Emax Immuno Assay system from Promega according to the manufacturer’s protocol (Madison, WI, USA). Absorbency was read in duplicate using a Bio-Rad Benchmark microplate reader (Hercules, CA, USA) at 450 nm. The coefficients of variance ranged between 2.9 and 8.1%. In total, 2498 (97.5%) of the subjects included in our sample had a BDNF value available. A previous NESDA paper3 (link) showed that anti-depressant-free currently depressed participants had lower BDNF levels than both HCs and their medicated currently depressed peers.
Verduijn J., Milaneschi Y., Schoevers R.A., van Hemert A.M., Beekman A.T, & Penninx B.W. (2015). Pathophysiology of major depressive disorder: mechanisms involved in etiology are not associated with clinical progression. Translational Psychiatry, 5(9), e649-.
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10

BDNF Quantification in Serum Samples

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Serum samples were kept frozen at −85℃ for a period between one and four years after which BDNF concentration (ng/mL) was measured using the Emax Immuno Assay system from Promega according to the manufacturer’s protocol. The undiluted serum was acid treated, which in a dilution-dependent way reliably increased the detectable BDNF. Subsequently, serum samples were diluted 100 times and stored again at −85℃ for BDNF assay the next day. After dilution, the BDNF concentrations were well within the range of the standard curve. The assay sensitivity threshold was ascertained at 1.56 ng/ml reflecting the minimum level of BDNF in the serum that could be reliably determined. Three samples were below this threshold and therefore excluded from all subsequent analyses. See our previous study37 (link) for more details on BDNF assessment.
Generaal E., Milaneschi Y., Jansen R., Elzinga B.M., Dekker J, & Penninx B.W. (2016). The brain-derived neurotrophic factor pathway, life stress, and chronic multi-site musculoskeletal pain. Molecular Pain, 12, 1744806916646783.
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