We performed an agarose gel retardation assay to determine the optimum pAuNS NPs/DNA-miRNAs ratio for miRs and pDNA co-loading on AuNS NPs. As per the aforementioned protocol, different NPs/DNA ratios of pAuNS complexes were prepared in nuclease-free DEPC water and then loaded in 0.7% agarose gel along with the equivalent amount of naked pcDNA as control and electrophoresed in Mini-Sub Cell GT Systems at 40 V for 45 min. After the run, we imaged the gel in the BioRad Gel Doc XR+ Gel Documentation system (Bio-Rad, Hercules, CA, USA) to further quantify and analyze the extent of DNA encapsulation. To visualize the miRNAs migration in gel electrophoresis, we used the Cy5 conjugated antimiR-21 for the encapsulation study. At the end of the run, the gel was imaged in a Lumina III In Vivo Imaging System (IVIS, PerkinElmer) using 2s acquisitions at filters compatible for Cy5 excitation and emission wavelengths.
Lumina 3 in vivo imaging system
Manufactured by PerkinElmer
Sourced in United States
The Lumina III in vivo Imaging System is a laboratory equipment product designed for non-invasive imaging of small animals. It provides advanced capabilities for visualizing and quantifying biological processes within living subjects.
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Lab products found in correlation
9 protocols using lumina 3 in vivo imaging system
1
Optimizing pAuNS NPs/DNA-miRNAs Ratio
2
Murine Tumor Xenograft and Metastasis Model
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C57BL/6 strain were used as experimental model to study effect of Ts-depletion in syngeneic LL/2 cells to prevent immune rejection.13 (link) Mice were anaesthetised using isoflurane and euthanised by cervical dislocation.
For subcutaneous injections, 1 × 106 cells resuspended in 50 µl 0.9% NaCl were mixed with Matrigel (Corning) in a ratio 1:1 (v:v). Cells were injected in right flanks of 10–15-weeks-old female C57BL/6, with eight mice per group. Calliper measurements were taken every 4th day and tumour volume was calculated using the formula (Length × Width2 × π)/6.
For tail-vein metastasis assay, 5 × 105 LL/2 pLKO and shTs cells were resuspended in 100 µl PBS and injected in the tail vein of female C57BL/6, with 10 mice per group. Lung metastases were monitored by bioluminescence imaging (BLI) 4 weeks after injection. Anesthetised mice were intraperitoneally injected with 150 mg/kg D-luciferin (Kayman Chemicals). Bioluminescence images were acquired with Lumina III in vivo Imaging System (IVIS, Perkin Elmer). For all the mice exposure time was maintained at 180 s. Raw IVIS images were analysed with Living Image software and the metastasis was represented as radiance.
For subcutaneous injections, 1 × 106 cells resuspended in 50 µl 0.9% NaCl were mixed with Matrigel (Corning) in a ratio 1:1 (v:v). Cells were injected in right flanks of 10–15-weeks-old female C57BL/6, with eight mice per group. Calliper measurements were taken every 4th day and tumour volume was calculated using the formula (Length × Width2 × π)/6.
For tail-vein metastasis assay, 5 × 105 LL/2 pLKO and shTs cells were resuspended in 100 µl PBS and injected in the tail vein of female C57BL/6, with 10 mice per group. Lung metastases were monitored by bioluminescence imaging (BLI) 4 weeks after injection. Anesthetised mice were intraperitoneally injected with 150 mg/kg D-luciferin (Kayman Chemicals). Bioluminescence images were acquired with Lumina III in vivo Imaging System (IVIS, Perkin Elmer). For all the mice exposure time was maintained at 180 s. Raw IVIS images were analysed with Living Image software and the metastasis was represented as radiance.
3
Biodistribution and In Vivo Imaging of BSA-Ce6@IrO2/MnO2 NPs
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For the biodistribution study, MDA-MB-231-bearing mice (n = 4) were intravenously injected with 200 μL of a suspension of BSA-Ce6@IrO2/MnO2 NPs in saline (3.0 mM Ir) at a dose of 10 mg/kg. The mice were sacrificed at different time points post-injection (2, 24, and 48 h) and the tumor tissue and major organs (liver, heart, lung, spleen, kidneys) were collected, weighed, and digested using aqua regia overnight. The Ir content of the samples was quantified using ICP-OES. To evaluate in vivo elimination, the feces of each mouse were collected, and each sample then dissolved in aqua regia. The amounts of Ir were analyzed through ICP-OES.
For in vivo fluorescence imaging, MDA-MB-231 tumor-bearing mice were randomly divided into three groups (n = 3) and given an intravenous injection of 200 μL of free Ce6 or BSA-Ce6@IrO2/MnO2 NPs (5 mg Ce6 equiv./kg) in saline via the tail vein. Fluorescence imaging was performed at different time intervals (1, 4, 8, 12, and 24 h post-injection) using a Lumina III in vivo imaging system (PerkinElmer, Waltham, MA, USA) with an excitation filter of 640 nm and an emission filter of 710 nm. 24 h after injection, the major organs (liver, spleen, kidney, heart, and lung) and tumor were excised for ex vivo imaging using the same instrument.
For in vivo fluorescence imaging, MDA-MB-231 tumor-bearing mice were randomly divided into three groups (n = 3) and given an intravenous injection of 200 μL of free Ce6 or BSA-Ce6@IrO2/MnO2 NPs (5 mg Ce6 equiv./kg) in saline via the tail vein. Fluorescence imaging was performed at different time intervals (1, 4, 8, 12, and 24 h post-injection) using a Lumina III in vivo imaging system (PerkinElmer, Waltham, MA, USA) with an excitation filter of 640 nm and an emission filter of 710 nm. 24 h after injection, the major organs (liver, spleen, kidney, heart, and lung) and tumor were excised for ex vivo imaging using the same instrument.
4
Xenograft Model for Doxorubicin Response
5
Tail Vein Metastasis Assay in Mice
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For tail vein metastasis assay, 1.5 × 106 cells were injected into tail vein of 6–8 weeks old female athymic nu/nu mice, with three mice per group. Lung metastases were monitored by bioluminescence imaging (BLI). Anesthetized mice were intraperitoneally injected with 200 mg/kg D-luciferin (Perkin Elmer). Bioluminescence images were acquired with Lumina III in vivo Imaging System (Perkin Elmer). Analysis was performed with live imaging software by measuring photon flux. Lungs were collected and fixed in 10% buffered formalin and processed to obtain paraffin blocks. Three-micron thick sections of formalin fixed paraffin embedded samples were stained using hematoxylin and eosin. Mice experiments were approved by Animal Ethics Committee of the Bilkent University.
6
Tumor Acidity Neutralization by SnSe Nanostructures
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To study the tumor acidity neutralization capacity of SnSe NSs, intratumoral pH values of CT26 tumorbearing mice before and at 24 h after intratumoral injection of SnSe NSs and inSnSe NSs (n = 3 per group, SnSe or inSnSe NSs = 4.5 mg/kg) were measured using an invasive pH microelectrode (PMHP5, PreSens, Regensburg, Germany). In addition, excised tumors with 1 mL PBS from different groups were prepared in tumor homogenate by a tissue homogenizer. The tumor homogenates untreated and treated with SnSe NSs incubated in buffers at a pH value of 6.5 were then mixed with the pH responsive fluorescence dye BCECF (10 μM, ApexBio) for 10 min before being imaged under a Lumina III in vivo imaging system (PerkinElmer, Waltham, MA, USA) (excitation = 440 and 480 nm, emission = 520 nm).
7
Fluorescence Imaging of DiI-labeled AFMSC
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Prior to AFMSC administration, we positioned anesthetized mice in an In Vivo Imaging System (IVIS) Lumina III (Perkin Elmer, MA, USA) and captured fluorescence at excitation 560 nm and emission 610 nm. We re-imaged the wounds on the mice immediately following administration of DiI-labeled AFMSCs. We continued imaging at 6, 12, 24 and 48 h post-administration and analyzed the data using Living Image software (Perkin Elmer, MA, USA). Mice were identified with numbers and treatments were blinded.
8
Bioluminescence Imaging of Cell Lines
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CAL-62 cells, CAL-62/Rluc cells, MDA-MB-231 and MDA-MB-231/Rluc (1.25 × 104, 2.5 × 104, 5 × 104, and 105 cells/well) were plated in white and clear-bottom 96-well plates with a serum-free DMEM medium. Twenty-four hours later, the appropriate substrate coelenterazine (Caliper, PerkinElmer) was added to each well. Rluc activity was determined by BLI, using the IVIS Lumina III instrument (In Vivo Imaging System, IVIS Lumina III, PerkinElmer).
9
Bioluminescence Imaging for Engraftment Evaluation
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The engraftment was evaluated through BLI, which was obtained using the In Vivo Imaging System (IVIS) Lumina III (Perkin Elmer, Santa Clara, CA, USA) and quantified by IVIS imaging software version 4.7.3 (Perkin Elmer, Santa Clara, CA, USA). Until the 40th day after transplantation, the images were captured weekly (4, 7, 19, 26, 33, and 40th day) and, after that, every three weeks (61, 82, 103, and 124th day). Right after the BMT, the animals were trichotomized to reduce the absorption of radiation by the fur melanin to mitigate the signal attenuation. For image acquisition, ten minutes prior to in vivo image, D-Luciferin 150 mg/kg i.p. (XenoLight, Perkin Elmer, Boston, MA, USA) was administered in host animals, this procedure was repeated each time point of evaluation described above. The images were obtained in dorsal, ventral, and lateral positions, using the following acquisition parameters: automatic exposure time, F/stop 4, binning 8, and FOV 12.9 cm.
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