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Samples were homogenized with a Bioruptor Next Gen (Diagenode) bath sonicator, set on HIGH and cooled to 4C, using 10 cycles of 10 s ON and 10 s OFF

and colleagues in the 1990s systematically examined various IGF-1 truncations and modifications, establishing that removal of the first three amino acids dramatically reduces IGFBP binding by 100-fold or more while maintaining or slightly increasing IGF-1 receptor affinity

The Journal of Biological Chemistry, 284, 2970429713

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Therefore, validation of results in primary cells is a fundamental step