Evaluate a novel product to simultaneously remove dead cells, cellular membranes,

描述(由申请人提供)：我们假设可以开发出一种具有商业竞争力的磁性纳米颗粒，以消除干细胞等细胞培养准备中的碎片。目前从健康培养物中去除多余物质的方法依赖于使用基于Annexin V的磁性颗粒的磁性负选择。膜联蛋白V与存在于死亡细胞和细胞膜碎片表面的磷脂磷脂酰丝氨酸(PS)结合。然后施加磁场，从健康的培养物中收集死亡细胞。尽管在商业上取得了成功，但基于Annexin V的技术有显著的缺点：(I)它仅限于去除含有PS的膜，(Ii)剥离的核和寡核苷酸不能被去除，(Iii)Annexin V不能高压灭菌，(Iv)Annexin V的活性会随着时间的推移而降低，(V)Annexin V需要含钙的缓冲液，(Vi)难以在血清中使用，(Vii)由于Annexin V的脱落而固有的不稳定，(Viii)可能具有高的内毒素水平，(Ix)制造和维护昂贵，因此大大增加了终端用户的成本。我们建议的产品将像基于Annexin V的磁性颗粒一样有效地消除培养物中的死亡细胞和细胞膜碎片。我们的产品还将(I)针对并消除核酸，(Ii)在缓冲液选择方面提供更大的灵活性，以及(Iii)耐高压灭菌以确保无菌。因为在某些情况下， 随着技术成熟到临床实践，细胞样本的制备将变得更加严格，这一应用的成功完成将对生物医学的许多领域产生深远和积极的影响，包括生殖医学、诊断应用和细胞治疗，如自体干细胞治疗。 与公共健康相关：该项目将评估一项从健康细胞培养物中去除不需要的有害碎片的新技术。由于细胞样本的制备条件将随着技术成熟进入临床实践而变得更加严格，我们的产品将对包括生殖医学、诊断应用和细胞治疗在内的许多生物医学领域产生深远和积极的影响。

DESCRIPTION (provided by applicant): We hypothesize that a commercially competitive magnetic nanoparticle can be developed to eliminate debris in the preparation of cell cultures, such as stem cells. Current methods to remove unwanted material from healthy cultures rely on magnetic negative selection using Annexin V-based magnetic particles. Annexin V binds to the phospholipid phosphatidylserine (PS) present on the surface of dead cells and cell membrane fragments. A magnetic field is then applied to collect dead cells from the healthy culture. Although commercially successful, Annexin V-based technology has significant drawbacks: (i) it is restricted to removing only PS-containing membranes, (ii) stripped nuclei and oligonucleotides cannot be removed, (iii) Annexin V cannot be autoclaved, (iv) Annexin V activity can degrade over time, (v) Annexin V requires calcium-containing buffers, (vi) is difficul to use in serum, (vii) is inherently unstable due to the sloughing of Annexin V, (viii) is likely t have high endotoxin levels, (ix) is expensive to manufacture and maintain, therefore adding significant cost to the end- user. Our proposed product will eliminate dead cells and cell membrane fragments from culture as effectively as Annexin V-based magnetic particles. Our product will also (i) target and eliminate nucleic acids, (ii) provide greater flexibility in buffe selection, and (iii) tolerate autoclaving to assure sterility. Because the conditions under which a cell sample is prepared will become more stringent as technologies mature into clinical practice, successful completion of this application will have a profound and positive impact on many sectors of biomedicine, including reproductive medicine, diagnostic applications and cell therapy, such as autologus stem cell therapy. PUBLIC HEALTH RELEVANCE: The project will evaluate a novel technology to remove unwanted and harmful debris from healthy cell cultures. Because the conditions under which a cell sample is prepared will become more stringent as technologies mature into clinical practice, our product will have a profound and positive impact on many sectors of biomedicine, including reproductive medicine, diagnostic applications and cell therapy.