RESUMEN
Immunogenic cell death (ICD) is a form of cell death that activates an adaptive immune response against dead-cell-associated antigens. Cancer cells killed via ICD can elicit antitumor immunity. ICD is efficiently induced by near-infrared photo-immunotherapy (NIR-PIT) that selectively kills target-cells on which antibody-photoabsorber conjugates bind and are activated by NIR light exposure. Advanced live cell microscopies showed that NIR-PIT caused rapid and irreversible damage to the cell membrane function leading to swelling and bursting, releasing intracellular components due to the influx of water into the cell. The process also induces relocation of ICD bio markers including calreticulin, Hsp70 and Hsp90 to the cell surface and the rapid release of immunogenic signals including ATP and HMGB1 followed by maturation of immature dendritic cells. Thus, NIR-PIT is a therapy that kills tumor cells by ICD, eliciting a host immune response against tumor.
Asunto(s)
Antineoplásicos Inmunológicos/farmacología , Cetuximab/farmacología , Citotoxicidad Inmunológica , Inmunoterapia/métodos , Rayos Infrarrojos , Neoplasias/terapia , Fármacos Fotosensibilizantes/farmacología , Fototerapia/métodos , Trastuzumab/farmacología , Escape del Tumor , Adenosina Trifosfato/metabolismo , Animales , Calreticulina/metabolismo , Línea Celular Tumoral , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Receptores ErbB/antagonistas & inhibidores , Receptores ErbB/inmunología , Femenino , Proteína HMGB1/metabolismo , Proteínas HSP70 de Choque Térmico/metabolismo , Proteínas HSP90 de Choque Térmico/metabolismo , Humanos , Ratones , Ratones Desnudos , Microscopía/métodos , Células 3T3 NIH , Neoplasias/inmunología , Neoplasias/metabolismo , Neoplasias/patología , Receptor ErbB-2/antagonistas & inhibidores , Receptor ErbB-2/genética , Receptor ErbB-2/inmunología , Factores de Tiempo , Transfección , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Tomographic phase microscopy (TPM) facilitates three-dimensional imaging of live cells based on quantitative measurement of the distribution of the refractive index, but without the need for specific staining. However, the limited imaging speed and the anisotropic resolution of the reconstructed refractive index map remain major obstacles to the extension and further application of TPM. To address these obstacles, we first formulate a general measurement model that linearises the relationship between the measurement data and refractive index map based on a system matrix. In this way, the measurement system is interpreted as a linear system in a complete manner. Then we propose a reconstruction framework for retrieving the refractive index map from the measurement data with reduced angular sample frequency and limited angular coverage of illumination. The framework aims to transform the reconstruction task into an optimisation scheme based on total variation norm regularisation, followed by an efficient solution using the accelerated alternating direction method of multipliers algorithm. Using this method, only sparse angular illuminations need to be collected, thus speeding up the imaging process. We obtained experimental results from both cell-mimic phantom data and real measurement data, which showed that the proposed method can improve the imaging speed while still providing refractive index images with better quality compared with a conventional reconstruction method.
Asunto(s)
Algoritmos , Imagenología Tridimensional/métodos , Microscopía de Contraste de Fase/métodos , Modelos Biológicos , Fantasmas de Imagen , Refractometría/métodos , Tomografía Computarizada por Rayos X/métodos , Humanos , IluminaciónRESUMEN
There is a need for a noninvasive technique to monitor living pluripotent stem cell condition without any labeling. We present an optical imaging technique that is able to capture information about optical path difference through the cell and cell adhesion properties simultaneously using a combination of quantitative phase microscopy (QPM) and interference reflection microscopy (IRM) techniques. As a novel application of QPM and IRM, this multimodal imaging technique demonstrated its ability to distinguish the undifferentiated status of human induced pluripotent stem (hiPS) cells quantitatively based on the variation of optical path difference between the nucleus and cytoplasm as well as hiPS cell-specific cell adhesion properties.
RESUMEN
The subject study demonstrates the imaging of cell activity by quantitatively assessing the motion of intracellular organelles and cell plasma membranes without any contrast agent. The low-coherent interferometric technique and phase-referenced phase shifting technique were integrated to reveal the depth-resolved distribution of intracellular motility. The transversal and vertical spatial resolutions were 0.56 µm and 0.93 µm, respectively, and the mechanical stability of the system was 1.2 nm. The motility of the cell was assessed by mean squared displacement (MSD) and we have compensated for the MSD by applying statistical noise analysis. Thus we show the significant change of intracellular motility after paraformaldehyde treatment in non-labeled cells.
Asunto(s)
Movimiento Celular , Imagenología Tridimensional/métodos , Microscopía de Contraste de Fase/métodos , Coloración y Etiquetado , Línea Celular Tumoral , Supervivencia Celular , Humanos , Factores de TiempoRESUMEN
We have developed a Linnik-type interference microscope provided with a low-coherent light source to obtain topographic images of an intact cellular membrane on a nanometer scale. Our technique is based on measurement of the interference between light reflected from the cell surface and a reference beam. The results show full field surface topography of cultured cells and reveal an intrinsic membrane motion of tens of nanometers.
Asunto(s)
Membrana Celular/ultraestructura , Aumento de la Imagen/instrumentación , Interferometría/instrumentación , Microscopía de Contraste de Fase/instrumentación , Nanotecnología/instrumentación , Tomografía de Coherencia Óptica/instrumentación , Tamaño de la Célula , Diseño de Equipo , Análisis de Falla de Equipo , Interferometría/métodos , Microscopía de Contraste de Fase/métodos , Sensibilidad y Especificidad , Tomografía de Coherencia Óptica/métodosRESUMEN
We report a Koehler-illumination-based full-field, actively stabilized, low-coherence phase-shifting interferometer, which is built on a white-light Michelson interferometer. By using a phase-stepping technique we can obtain full-field phase images of the sample. An actively stabilized phase-lock circuit is employed in the system to reduce phase noise. An application to human epithelial cells (HeLa cells) is achieved in our experiment. The advancement of this technique rests in its ability to take images of unstained biological samples quantitatively and on a nanometer scale.
Asunto(s)
Aumento de la Imagen/instrumentación , Interferometría/instrumentación , Microscopía de Contraste de Fase/instrumentación , Transductores , Diseño de Equipo , Análisis de Falla de Equipo , Retroalimentación , Células HeLa , Humanos , Aumento de la Imagen/métodos , Interpretación de Imagen Asistida por Computador/métodos , Interferometría/métodos , Luz , Microscopía de Contraste de Fase/métodos , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
By use of the Fourier decomposition of a low-coherence optical image field into two spatial components that can be controllably shifted in phase with respect to each other, a new high-transverse-resolution quantitative-phase microscope has been developed. The technique transforms a typical optical microscope into a quantitative-phase microscope, with high accuracy and a path-length sensitivity of lambda/5500, which is stable over several hours. The results obtained on epithelial and red blood cells demonstrate the potential of this instrument for quantitative investigation of the structure and dynamics associated with biological systems without sample preparation.
Asunto(s)
Eritrocitos/citología , Microscopía de Contraste de Fase/instrumentación , Microscopía de Contraste de Fase/métodos , Tomografía de Coherencia Óptica/instrumentación , Tomografía de Coherencia Óptica/métodos , Células Cultivadas , Análisis de Falla de Equipo , Análisis de Fourier , Células HeLa , Humanos , Fantasmas de Imagen , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
We describe a quantitative phase-imaging interferometer in which phase shifting and noise cancellation are performed by an active feedback loop using a reference laser. Depth gating via low-coherence light allows phase measurement from weakly reflecting biological samples. We demonstrate phase images from a test structure and living cells.