Park Jason Y, Gunpat Joshua, Liu Li, Edwards Brooks, Christie Alana, Xie Xian-Jin, Kricka Larry J, Mason Ralph P
Department of Pathology, University of Texas Southwestern Medical Center and Children's Medical Center, Dallas, TX, USA; Eugene McDermott Center for Human Growth and Development, University of Texas Southwestern Medical Center, Dallas, TX, USA.
Luminescence. 2014 Sep;29(6):553-8. doi: 10.1002/bio.2666. Epub 2014 Apr 24.
Commercial chemiluminescent reagents emit across a broad portion of the electromagnetic spectrum (400-500 nm). A challenge to the use of chemiluminescence to monitor biological processes is the presence of interfering substances in the biological optical window. In the present study, longer wavelength emitting fluorophores (the organic dyes Alexa 568 and Alexa 647), and a semiconductor nanoparticle (QDOT800) were used to red-shift the emission from commercially available 1,2-dioxetane-based chemiluminescent substrate reactions. By adding non-conjugated fluorescent emitters into chemiluminescent reaction mixtures, an emission peak occurred at the predicted wavelength of the fluorescent emitter. The excitation and emission from QDOT800 was preserved in the presence of a 100 µm-thick glass barrier separating it from the chemiluminescent reaction components. The maximum tissue phantom penetration by QDOT800 emission was 8.5 mm; in comparison, the native chemiluminescent emission at 500 nm was unable to penetrate the thinnest tissue phantom of 2.5 mm. The described method for red-shifted emissions from chemiluminescent reactions does not require direct interaction between the chemiluminescent reaction and the fluorescent emitters. This suggests that the mechanism of chemiluminescent excitation of fluorophores and QDOT800 is not exclusive to chemiluminescence resonance energy transfer or sensitized chemiluminescence, but rather by broad energization from the native chemiluminescent emission.
商业化学发光试剂在电磁光谱的广泛区域（400 - 500纳米）发射光。利用化学发光监测生物过程面临的一个挑战是生物光学窗口中存在干扰物质。在本研究中，使用发射更长波长的荧光团（有机染料Alexa 568和Alexa 647）以及一种半导体纳米颗粒（QDOT800）来使基于1,2 - 二氧杂环丁烷的市售化学发光底物反应的发射光发生红移。通过向化学发光反应混合物中添加非共轭荧光发射体，在荧光发射体的预测波长处出现了一个发射峰。在存在将QDOT800与化学发光反应组分隔开的100微米厚玻璃屏障的情况下，QDOT800的激发和发射得以保留。QDOT800发射光在组织模型中的最大穿透深度为8.5毫米；相比之下，500纳米处的天然化学发光发射无法穿透最薄的2.5毫米组织模型。所描述的使化学发光反应发射光发生红移的方法并不要求化学发光反应与荧光发射体之间直接相互作用。这表明荧光团和QDOT800的化学发光激发机制并非仅限于化学发光共振能量转移或敏化化学发光，而是由天然化学发光发射的广泛能量激发所致。
