Although it is passively released from the cells unmodified, Cyt C can still be an apoptosis marker as Cyt C is released from the mitochondrial intermembrane space into the cytosol in the apoptosis-specific process. the medium before plasma membrane integrity was compromised. == Conclusions == Taken together, these data suggest that serum fortilin levels reflect the degree and extent of apoptosis occurring in vivo. == General significance == Fortilin is a viable serum biomarker of in vivo apoptosis and can be utilized to noninvasively assess the status of in vivo apoptosis in humans. Keywords:Apoptosis, Biomarker, Fortilin, Programmed cell death == Graphical abstract == == Highlights == Ultra-sensitive fortilin ELISA has been developed. Fortilin circulates in blood. Fortilin serum levels become highly elevated after apoptosis-inducing therapy. Fortilin is more robust and sensitive than other serum apoptosis markers. Fortilin is actively secreted before plasma membrane becomes disrupted. Fortilin is an excellent serum biomarker of in vivo apoptosis. == 1. Introduction == Approximately 50 to 70 billion cells undergo Akt-l-1 apoptosis each day in the average normal adult[1]. Serum biomarkers of apoptosis molecules that can be readily and objectively measured as indicators of normally and pathologically occurring apoptosis at tissue and organ levels would allow clinicians to Akt-l-1 easily monitor the status of apoptosis associated with the diseases and conditions they treatsuch as apoptosis-induced skeletal muscle atrophy resulting from cancer (cachexia), aging (sarcopenia)[2], starvation, denervation, disuse, and inflammation[3]. Cancer cells undergo apoptosis at a higher rate than do normal cells and massively apoptose in response to radiation therapy and chemotherapy[4]. Serum biomarkers of apoptosis could thus allow clinicians to screen patients for certain cancers or to monitor the response of patients with cancer to anti-cancer chemo- or radiation therapy[5]. Thus far, three serum biomarkers of apoptosis have been reported in the literature including the fragmented cytokeratin-18 (fCK18, detectable by the M30 antibody), nucleosomally-cleaved genomic DNA (n-DNA), and cytochrome c (Cyt C)[5]each with notable KRT19 antibody limitations to their utility. The utility of fCK18 is limited to apoptosis occurring in cells of epithelial origin[6]. The utility of circulating n-DNA is diminished because it can be rapidly degraded by serum DNases[7]. Cyt C is reportedly Akt-l-1 released from both apoptotic[8]and necrotic cells[9], depending on the extent of cellular damage, thus Akt-l-1 limiting its specificity. Further, these candidate serum apoptosis biomarkers have not been extensively characterized or validated at clinical, whole animal, and cellular levels. Fortilin, also known as translationally controlled tumor protein (TCTP) and IgE-dependent histamine releasing factor (HRF), is a 172-amino acid nuclear-cytosolic shuttle protein that was originally cloned in 1989 by Gross and others as a molecule abundantly expressed in tumor cells[10]. A multifunctional protein implicated in various cellular functions[11],[12],[13],[14],[15],[16], fortilin possesses potent anti-apoptotic activity[12],[17],[18],[19],[20],[21],[22]. It binds the sequence-specific DNA binding domain of p53 and prevents p53 from transcriptionally activating the proapoptotic gene Bax[23]. Fortilin also binds to and stabilizes MCL1[22], a macrophage survival factor[24],[25]. In addition to being intracellularly located, fortilin can be trafficked into exosomes small secretory vesicles and eventually be released into the extracellular space in an ER/Golgi-independent fashion[15]. However, it remains unknown if fortilin actually circulates in the blood of normal humans and animals. Further, it is not known if serum fortilin levels change in response to various pathophysiological conditions, including apoptosis occurring in body tissues. Since it is a potent anti-apoptotic molecule that can be secreted into the extracellular space, we hypothesized that fortilin could be a viable serum apoptosis biomarker..