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Softmax pro acquisition and analysis software

Manufactured by Molecular Devices
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SoftMax Pro Acquisition and Analysis software is a comprehensive data acquisition and analysis platform for microplate readers. It is designed to streamline the process of collecting, managing, and interpreting data from various types of microplate-based experiments.

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Lab products found in correlation

Spectramax m5 microplate reader, by Molecular Devices (5 mentions) Anti mouse il 10, by R&D Systems (2 mentions) Mouse anti il12, by BD (2 mentions) Phosphatase conjugated streptavidin, by Southern Biotech (2 mentions) P nitrophenyl phosphatase pnpp substrate, by Southern Biotech (2 mentions) Ct dna, by Merck Group (1 mentions) Anti tnf α, by R&D Systems (1 mentions) Anti m csf, by R&D Systems (1 mentions) Anti il 10, by R&D Systems (1 mentions) Anti il 6, by R&D Systems (1 mentions) Prestoblue reagent, by Thermo Fisher Scientific (1 mentions) Cellrox green reagent, by Thermo Fisher Scientific (1 mentions)

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SoftMax® Pro Microplate Data Acquisition and Analysis software SoftMax Pro Data Acquisition and Analysis Software SoftMax Pro software SoftMax Pro SoftMax software

5 protocols using softmax pro acquisition and analysis software

1

Quantifying Cytokine Levels via ELISA

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ELISA was performed based on the protocol described previously10 (link). Immunol 2HB-microtiter plates were coated with mouse anti IL-10 (R&D Systems) or mouse anti-IL12 (BD Biosciences) followed by blocking with 2% BSA-PBS for 2 hr. Culture supernatants or diluted sera were added after washing and incubated overnight at 4 °C. Secondary Ab labeled with biotin were added to plates, and incubated for 2 hr. Next plates were developed by phosphatase-conjugated streptavidin (AKP) followed by the addition of p-nitrophenyl phosphatase (pNPP) substrate (Southern Biotech). Optical density was measured using a SpectraMax M5 microplate Reader and SoftMax Pro Acquisition and Analysis software (Molecular Devices).
Horuluoglu B.H., Kayraklioglu N., Tross D, & Klinman D. (2020). PAM3 protects against DSS-induced colitis by altering the M2:M1 ratio. Scientific Reports, 10, 6078.
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2

Quantification of Anti-Cytokine Antibodies

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Immunol 2HB-microtiter plates were coated with human anti-TNFα (R&D Systems), mouse anti-IL12 (BD Biosciences), mouse anti-TNFα (R&D Systems) or mouse anti-IL-10 (R&D Systems) and then blocked with 2% BSA-PBS for 2 hr. After washing, culture supernatants or serially diluted sera were added overnight at 4° C. Plates were washed, incubated with biotin-labeled secondary Ab and developed by adding phosphatase-conjugated streptavidin (AKP) followed by p-nitrophenyl phosphatase (pNPP) substrate (Southern Biotech). Optical density was measured using a SpectraMax M5 microplate Reader and SoftMax Pro Acquisition and Analysis software (Molecular Devices).
IgG anti-dsDNA was detected by coating Immunol 2HB-microtiter plates with 4 μg/ml of CT-DNA (Sigma) in DNA binding solution (Abcam) for 4 hr. The plates were blocked with 2% BSA-PBS and incubated with diluted mouse serum as above. Bound Ab was detected with biotin labeled IgG (Invitrogen) followed by phosphate-conjugated streptavidin and pNPP substrate.
Horuluoglu B., Bayik D., Kayraklioglu N., Goguet E., Kaplan M.J, & Klinman D.M. (2019). PAM3 supports the generation of M2-like macrophages from lupus patient monocytes and improves disease outcome in murine lupus. Journal of autoimmunity, 99, 24-32.
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3

Quantitative Cytokine Profiling from Cell Cultures

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Cell supernatants were collected on day 3 and frozen until further use. Immunol 2HB microtiter plates (Thermo Scientific) were coated with anti-cytokine antibodies anti-IL-6 (Clone #6708), anti-IL-10 (Clone #127107), anti-TNFα (Clone #28401), and anti-M-CSF (Clone #21113) (R&D Systems, Minneapolis, MN, USA) and then blocked with PBS/2% BSA. Serially diluted standards and culture supernatants were added to these plates overnight. Plates were incubated with biotinylated anti-cytokine Ab (R&D Systems), followed by phosphatase-streptavidin (BD Biosciences) and K-Gold PNPP Substrate (Neogen Corporation, Lexington, KY, USA). Human IL-12p70 Quantikine, IL-4 Quantikine, and TGFβ1 Quantikine ELISAs were performed based on manufacturer’s instructions (R&D Systems). ELISAs were read using a SpectraMax M5 Microplate Reader and SoftMax Pro Acquisition and Analysis Software (both Molecular Devices, Sunnyvale, CA, USA).
Bayik D., Tross D, & Klinman D.M. (2018). Factors Influencing the Differentiation of Human Monocytic Myeloid-Derived Suppressor Cells Into Inflammatory Macrophages. Frontiers in Immunology, 9, 608.
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4

PrestoBlue Viability Assay Protocol

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Cell viability was assessed using the PrestoBlue™ reagent (Thermo Inc., Waltham, MA). PrestoBlue is a soluble tetrazolium salt that readily enters cells, where it is reduced, in metabolically active (viable) cells, by mitochondrial dehydrogenases and reductases, to an insoluble, blue-colored, red-fluorescent formazan product. The PrestoBlue™ viability assay was performed after 24 h exposure to digestas, using 96-well plate co-cultures, according to manufacturer’s instructions. Briefly, treated and negative control (100% viable) untreated cells were washed 3 times with 200 μL/well PBS, and 100 μL of 10% PrestoBlue reagent was added to each well. Plates were then incubated at 37 ºC for 15 minutes, and fluorescence was measured at 560 nm (excitation)/590 nm (emission) using a SpectraMax M-5 microplate reader and SoftMax Pro acquisition and analysis software (Molecular Devices, San Jose, CA).
DeLoid G.M., Cao X., Bitounis D., Singh D., Llopis P.M., Buckley B, & Demokritou P. (2021). Toxicity, uptake, and nuclear translocation of ingested micro-nanoplastics in an in vitro model of the small intestinal epithelium. Food and chemical toxicology : an international journal published for the British Industrial Biological Research Association, 158, 112609.
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5

Measurement of Oxidative Stress in Co-Cultures

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Oxidative stress was assessed by measuring cellular reactive oxygen species (ROS) accumulation after 6 h exposure to digestas in 96-well co-cultures. ROS production (oxidative stress) was assessed using the CellROX green reagent (Thermo Inc., Waltham, MA) according to the manufacturer’s instructions. Briefly, a 5 mM working solution of the CellROX green reagent was prepared from 20 mM stock by diluting in glucose-free DMEM media without FBS. Media was removed from test wells and replaced with 100 μL of working solution, and plates were incubated for 30 minutes at 37 ºC. Cells were then washed 3 times with 200 μL/well PBS, and fluorescence was measured at 480 nm (excitation)/520 nm (emission) using a SpectraMax M-5 microplate reader and SoftMax Pro acquisition and analysis software (Molecular Devices, San Jose, CA), quantifying the amount of oxidized CellROX green reagent.
DeLoid G.M., Cao X., Bitounis D., Singh D., Llopis P.M., Buckley B, & Demokritou P. (2021). Toxicity, uptake, and nuclear translocation of ingested micro-nanoplastics in an in vitro model of the small intestinal epithelium. Food and chemical toxicology : an international journal published for the British Industrial Biological Research Association, 158, 112609.
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