Biophotonics Solutions for Quantitative Live Cell Near Infrared Raman Spectral Analysis of DNA Damage in Spermatozoa

用于对精子 DNA 损伤进行活细胞近红外拉曼光谱定量分析的生物光子学解决方案

• 批准号: EP/C54711X/1
• 负责人: Alistair Elfick
• 金额: $ 25.43万
• 依托单位: University of Edinburgh
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2006
• 资助国家: 英国
• 起止时间: 2006 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=EP%2FC54711X%2F1
• 关键词: Biophotonics; Solutions; Quantitative; Live; Cell

Infertility affects at least one in six couples in the UK. The most common cause man infertility. Male factor infertility is usually characterised by sperm which are too weak to swim to the egg or don't swim at all. The genetic material (DNA) held in these unhealthy sperm is highly likely to be damaged.A treatment which is used in these cases is to inject the sperm into the egg in the laboratory. However, there is at present no way to ensure that the DNA which is transferred in this process is of good quality. There is concern in the medical community that this may lead to high rates of disease such as childhood cancer for children conceived by sperm injection.All the currently available tests for DNA damage require the sperm to be broken open and the DNA extracted to be chemically analysed. The proposed work will develop a method to trap a single sperm and measure its DNA integrity without hurting the sperm. A good sperm can then be selected to use to fertilise the egg. The technique developed should greatly reduce the risk of abnormalities in the children conceived by sperm injection. It may also increase the success rates for assisted fertilisation.We propose to combine two methods which use laser light: optical tweezers and Raman spectroscopy. If a beam of laser light is focused very tightly it can create enough force to hold a very small object - this is called optical trapping or optical tweezers. The sperm which may be trying to swim away will be held where we want it by optical trapping. Now we have trapped our very small and wriggling cell we have to characterise its DNA. This will be done by using a different colour of laser to vibrate the molecules of the cell. We can then look at how the vibrations affect the light. By using powerful statistical analysis we can build up knowledge of how damage DNA vibrates compared to good DNA.The work will involve a lot of optical design to create the instrument needed to do this work. We will then have to prove that our lasers themselves are not causing any additional damage to, the sperm. Tweezers and Raman have both been used before on cells without affecting them but we need to be absolutely sure that this is the case for sperm too. Next we'll build statistical models by taking healthy sperm and producing a known level of damage in them with chemicals and teaching our Raman software what that sperm 'looks' like. Finally, we'll take sperm from infertile men which have been sorted into different degrees of healthiness and we'll see if our instrument can tell the difference between them.There is a good chance that this instrument will be able to be used in laboratories around the world which are treating people with infertility or doing research into reproduction.

在英国，不孕症至少影响六分之一的夫妇。最常见的原因是男性不育。男性因素不育的特点通常是精子太弱，无法游向卵子或根本不游泳。这些不健康精子中的遗传物质（DNA）极有可能受到损害。在这些情况下使用的治疗方法是在实验室中将精子注射到卵子中。然而，目前没有办法确保在这一过程中转移的DNA是高质量的。医学界担心，这可能会导致高发病率，例如通过精子注射怀孕的儿童患上儿童癌症，目前所有可用的DNA损伤测试都需要将精子破碎，并提取DNA进行化学分析。这项拟议中的工作将开发一种方法来捕获单个精子，并在不伤害精子的情况下测量其DNA完整性。然后可以选择一个好的精子来使卵子受精。这项技术的发展将大大降低通过精子注射受孕的孩子畸形的风险。我们建议结合联合收割机和拉曼光谱两种使用激光的方法。如果一束激光非常紧密地聚焦，它可以产生足够的力来保持一个非常小的物体-这被称为光阱或光镊。可能试图游走的精子将被光学捕获器固定在我们想要的地方。现在我们已经捕获了我们非常小的蠕动细胞，我们必须复制它的DNA。这将通过使用不同颜色的激光来振动细胞的分子来完成。然后我们可以看看振动是如何影响光的。通过使用强大的统计分析，我们可以建立受损DNA与正常DNA相比如何振动的知识。这项工作将涉及大量的光学设计，以创建完成这项工作所需的仪器。然后，我们将不得不证明我们的激光本身不会对精子造成任何额外的伤害。镊子和拉曼都曾在细胞上使用过而没有影响它们，但我们需要绝对肯定精子也是如此。接下来，我们将建立统计模型，方法是取健康的精子，用化学物质在它们体内产生已知水平的损伤，并教会我们的拉曼软件精子的“样子”。最后，我们将从不育男性中提取精子，这些精子被分为不同的健康程度，我们将看看我们的仪器是否能区分它们之间的区别。这台仪器很有可能能够在世界各地的实验室中使用，这些实验室正在治疗不育症或进行生殖研究。

项目成果

Off Axis Fresnel Zone Plates for White Light 3D Microscopy

用于白光 3D 显微镜的离轴菲涅尔波带板

• DOI: 10.1364/fio.2006.ftuw3
• 发表时间: 2006
• 作者: Raheem R

The changing biochemical composition and organisation of the murine oocyte and early embryo as revealed by Raman spectroscopic mapping

• DOI: 10.1002/jrs.2964
• 发表时间: 2012-01-01
• 期刊: JOURNAL OF RAMAN SPECTROSCOPY
• 影响因子: 2.5
• 作者: Davidson, Bryony;Spears, Norah;Elfick, Alistair
• 通讯作者: Elfick, Alistair

Raman micro-spectroscopy can be used to investigate the developmental stage of the mouse oocyte.

• DOI: 10.1371/journal.pone.0067972
• 发表时间: 2013
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Davidson B;Murray AA;Elfick A;Spears N
• 通讯作者: Spears N

Label-Free Analysis of Oocyte Development Using Raman Micro-Spectroscopy

使用显微拉曼光谱对卵母细胞发育进行无标记分析

• 发表时间: 2009
• 作者: B Davidson

3D Optical trapping Design for Multi-Purpose Microscopes with Space Constraints

适用于空间受限的多功能显微镜的 3D 光学捕获设计

• 发表时间: 2008
• 作者: R Raheem