Serum HGF concentrations were determined with the commercially available enzyme‐linked immunosorbent assay kit HGF ELISA kit (Thermo Fisher, Waltham, USA), and the assays were performed, according to the manufacturer’s instructions. The optical readout was conducted with the microplate reader Infinite M200 and software magellan version 7.2 (Tecan, Männedorf, Switzerland).
Magellan version 7
Manufactured by Tecan
Sourced in Switzerland
Magellan version 7.1 is a software package for the control and data analysis of Tecan's microplate readers. It provides a comprehensive set of tools for instrument operation, data acquisition, and data processing. The software supports a wide range of Tecan's microplate reader models and offers flexible configuration options to meet the needs of diverse laboratory applications.
Automatically generated - may contain errors
7 protocols using magellan version 7
1
Serum HGF Quantification by ELISA
2
Quantifying sMET Levels via ELISA
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sMET concentrations were determined with the enzyme‐linked immunosorbent assay kit (soluble) cMet ELISA Kit (Thermo Fisher, Waltham, MA, USA), and the assays were performed according to manufacturer's instructions. The final optical readout was conducted with the microplate reader Infinite M200 and software magellan version 7.2 (Tecan, Männedorf, Switzerland).
3
Quantification of Serum IL-7 Levels
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Serum of study participants was stored at -80 °C for concomitant IL-7 measurement at the University Childrens Hospital in Duesseldorf. Serum IL-7 concentrations were measured as described previously [16, 20] . In brief, we used a commercially available highly sensitive ELISA (Human IL-7 Quantikine HS ELISA kit, R&D Systems) according to manufacturer's instructions. Because of low IL-7 serum concentrations, we included standards on each plate to increase the accuracy of these assays. All samples were measured at least as duplicates. Optical density was determined using an ELISA reader (Infinite M200, Tecan). IL-7 concentrations were calculated from the standard curve by applying 4-parametric logistic regression (Magellan version 7.2, Tecan).
4
Automated Liquid Handling for Spectrophotometry
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Liquid handling studies were carried out using a Freedom EVO® 200 LHS from Tecan (Crailsheim, Germany). The LHS was equipped with one robotic manipulator arm (ROMA) and one liquid handling arm (LIHA) with eight independently controllable pipetting needles. Liquid pipetting was executed with stainless steel standard tips. Absorbance of elution fractions was measured with an Infinite M200 microplate reader from Tecan. Experimental LHS protocols including automated photospectrometry were developed using EVOware® 2.5 and Magellan (version 7.1) from Tecan. Additionally, a TE‐Link (Tecan) was installed for transferring elution plates under the column array carrier to collect fractions. Plates were stored in hotels. Deep well plates (DWPs), used for storing elution buffers, and ultraviolet (UV) microtiter plates (MTPs) containing elution fractions were from Greiner (Frickenhausen, Germany). All MTPs were covered with a lid to minimize evaporation. Small fractions with less than 100 µL volume were collected in half‐area UV plates. The size of dilutor syringes was 1 mL, enabling volume pipetting in the range of 1–1000 µL. Liquid volume application greater than 1 mL was conducted via iterative loops.
5
Estradiol-17ß ELISA Quantification
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For estradiol-17ß analysis a commercial estradiol-17ß ELISA Kit (IBL International, Hamburg, Germany) was used. Assays were conducted following the manufacturer’s instructions and samples were analyzed in duplicates. The kit used detects the entire E2. The absorption of the plate was detected with an ELISA microplate reader (Tecan, Crailsheim, Germany) at 450 nm with a reference wavelength at 620 nm.
The concentrations and coefficients of variation of the assays were calculated using the microplate reader software (Magellan® Version 7.1, Tecan Austria, Salzburg, Austria). For estradiol the intraassay coefficient of variation was 6% and the interassay coefficient of variation was 9%.
The concentrations and coefficients of variation of the assays were calculated using the microplate reader software (Magellan® Version 7.1, Tecan Austria, Salzburg, Austria). For estradiol the intraassay coefficient of variation was 6% and the interassay coefficient of variation was 9%.
6
suPAR Measurement by ELISA
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For suPAR analysis, enzyme-linked immunosorbent assay (suPARnostic® AUTO Flex ELISA, ViroGates, Birkerod, Denmark) was performed according to manufacturer’s instructions [11 (link)]. Briefly, serum samples diluted 1:10 and peroxidase-conjugated anti-suPAR was mixed and then incubated in an anti-suPAR pre-coated 96-well plate. After 1 h incubation, tetramethylbenzidine substrate was added and the enzymatic reaction was stopped after 20 min by adding 2 N sulfuric acid and read at 450 nm (plate reader Sunrise, Tecan, Männedorf, Switzerland; software Magellan version 7.1, Tecan). All samples were run in duplicates.
A large Danish study of 5 538 adult individuals showed a mean serum concentration of suPAR of 3.51 ng/mL for men and 3.90 ng/mL for women [16 (link)]. The highest value among the controls herein (> 3.6 ng/mL) was used as cut-off; this corresponded to the 98th percentile among the healthy controls.
A large Danish study of 5 538 adult individuals showed a mean serum concentration of suPAR of 3.51 ng/mL for men and 3.90 ng/mL for women [16 (link)]. The highest value among the controls herein (> 3.6 ng/mL) was used as cut-off; this corresponded to the 98th percentile among the healthy controls.
7
Quantitative ELISA for Osteopontin in SLE
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