Dataset Information

Visible and near infrared fluorescence spectral flow cytometry.

ABSTRACT: There is a long standing interest in measuring complete emission spectra from individual cells in flow cytometry. We have developed flow cytometry instruments and analysis approaches to enable this to be done routinely and robustly. Our spectral flow cytometers use a holographic grating to disperse light from single cells onto a CCD for high speed, wavelength-resolved detection. Customized software allows the single cell spectral data to be displayed and analyzed to produce new spectra-derived parameters. We show that familiar reference and calibration beads can be employed to quantitatively assess instrument performance. We use microspheres stained with six different quantum dots to compare a virtual bandpass filter approach with classic least squares (CLS) spectral unmixing, and then use antibody capture beads and CLS unmixing to demonstrate immunophenotyping of peripheral blood mononuclear cells using spectral flow cytometry. Finally, we characterize and evaluate several near infrared (NIR) emitting fluorophores for use in spectral flow cytometry. Spectral flow cytometry offers a number of attractive features for single cell analysis, including a simplified optical path, high spectral resolution, and streamlined approaches to quantitative multiparameter measurements. The availability of robust instrumentation, software, and analysis approaches will facilitate the development of spectral flow cytometry applications.

SUBMITTER: Nolan JP

PROVIDER: S-EPMC3594514 | biostudies-literature | 2013 Mar

REPOSITORIES: biostudies-literature

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Publications

Visible and near infrared fluorescence spectral flow cytometry.

Nolan John P JP Condello Danilo D Duggan Erika E Naivar Mark M Novo David D

Cytometry. Part A : the journal of the International Society for Analytical Cytology 20121206 3

There is a long standing interest in measuring complete emission spectra from individual cells in flow cytometry. We have developed flow cytometry instruments and analysis approaches to enable this to be done routinely and robustly. Our spectral flow cytometers use a holographic grating to disperse light from single cells onto a CCD for high speed, wavelength-resolved detection. Customized software allows the single cell spectral data to be displayed and analyzed to produce new spectra-derived p ...[more]

PMID: 23225549

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Project description:Quartz is the most abundant mineral on the earth's surface. It is spectrally active in the longwave infrared (LWIR) region with no significant spectral features in the optical domain, i.e., visible-near-infrared-shortwave-infrared (Vis-NIR-SWIR) region. Several space agencies are planning to mount optical image spectrometers in space, with one of their missions being to map raw materials. However, these sensors are active across the optical region, making the spectral identification of quartz mineral problematic. This study demonstrates that indirect relationships between the optical and LWIR regions (where quartz is spectrally dominant) can be used to assess quartz content spectrally using solely the optical region. To achieve this, we made use of the legacy Israeli soil spectral library, which characterizes arid and semiarid soils through comprehensive chemical and mineral analyses along with spectral measurements across the Vis-NIR-SWIR region (reflectance) and LWIR region (emissivity). Recently, a Soil Quartz Clay Mineral Index (SQCMI) was developed using mineral-related emissivity features to determine the content of quartz, relative to clay minerals, in the soil. The SQCMI was highly and significantly correlated with the Vis-NIR-SWIR spectral region (R2 = 0.82, root mean square error (RMSE) = 0.01, ratio of performance to deviation (RPD) = 2.34), whereas direct estimation of the quartz content using a gradient-boosting algorithm against the Vis-NIR-SWIR region provided poor results (R2 = 0.45, RMSE = 15.63, RPD = 1.32). Moreover, estimation of the SQCMI value was even more accurate when only the 2000-2450 nm spectral range (atmospheric window) was used (R2 = 0.9, RMSE = 0.005, RPD = 1.95). These results suggest that reflectance data across the 2000-2450 nm spectral region can be used to estimate quartz content, relative to clay minerals in the soil satisfactorily using hyperspectral remote sensing means.

Dataset Information

Visible and near infrared fluorescence spectral flow cytometry.

Publications

Visible and near infrared fluorescence spectral flow cytometry.

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