Immunodepletion of EGF from breast milk was performed as in 21 (link). Briefly, 500ul aliquots of freshly harvested murine breast milk were filtered via Amicon Ultra 10K Centrifugal Filters (Millipore) at 13,000 RPM for 15 minutes. Filtered aliquots were then incubated at 4 degrees Celsius for 30 minutes with control goat IgG antibody (10 μl) and protein A/G-agarose (20 μl, Santa Cruz Biotechnology), and centrifuged at 3000RPM for 30 seconds. The supernatant was incubated for 1 h at 4 degrees Celsius with 2μg of anti-EGF antibody (Santa Cruz Biotechnology) followed by overnight incubation on a rotator with an additional 20 μl of protein A/G-agarose. Supernatants were collected after centrifugation at 3000RPM for 30 seconds and an EGF ELISA (Abcam) was performed as per manufacturers instructions using a 1:500 dilution (wild-type murine breast milk [22907±976 pg/ml]; EGF-depleted murine breast milk 468.5±87.1 pg/ml].
Egf elisa
Manufactured by Abcam
The EGF ELISA is a quantitative sandwich enzyme-linked immunosorbent assay designed to measure the concentration of epidermal growth factor (EGF) in biological samples. It is a reliable and sensitive tool for researchers to investigate EGF levels in their studies.
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2 protocols using egf elisa
1
Immunodepletion of EGF from Murine Breast Milk
2
Immunodepletion of EGF from Murine Breast Milk
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Immunodepletion of EGF from breast milk was performed as in 21 (link). Briefly, 500ul aliquots of freshly harvested murine breast milk were filtered via Amicon Ultra 10K Centrifugal Filters (Millipore) at 13,000 RPM for 15 minutes. Filtered aliquots were then incubated at 4 degrees Celsius for 30 minutes with control goat IgG antibody (10 μl) and protein A/G-agarose (20 μl, Santa Cruz Biotechnology), and centrifuged at 3000RPM for 30 seconds. The supernatant was incubated for 1 h at 4 degrees Celsius with 2μg of anti-EGF antibody (Santa Cruz Biotechnology) followed by overnight incubation on a rotator with an additional 20 μl of protein A/G-agarose. Supernatants were collected after centrifugation at 3000RPM for 30 seconds and an EGF ELISA (Abcam) was performed as per manufacturers instructions using a 1:500 dilution (wild-type murine breast milk [22907±976 pg/ml]; EGF-depleted murine breast milk 468.5±87.1 pg/ml].
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