Hybrid reconstruction framework for model-based multispectral bioluminescence tomography based on Alpha-divergence

Bioluminescence tomography(BLT)is a promising imaging modality that can provide noninvasive three-dimensional visualization information on tumor distribution.In BLT reconstruction,the widely used methods based on regularization or greedy strategy face problems such as over-sparsity,over-smoothing,spatial discontinuity,poor robustness,and poor multi-target resolution.To deal with these problems,combining the advantages of the greedy strategies as well as regularization methods,we propose a hybrid reconstruction framework for model-based multispectral BLT using the support set of a greedy strategy as a feasible region and the Alpha-divergence to combine the weighted solutions obtained by L1-norm and L2-norm regularization methods.In numerical simulations with digital mouse and in vivo experiments,the results show that the proposed framework has better localization accuracy,spatial resolution,and multi-target resolution.

Study on Shigella Detection by ATP Bioluminescence Magnetic Enzyme Immunoassay

[Objective] The research aimed to establish a rapid detection method for Shigella. [Method] Combining immunomagnetic separation technology with ATP bioluminescence technology, a new kind of fast and accurate ATP bioluminescence magnetic enzyme immunoassay technique for Shigella was established. [Result] Using ATP bioluminescence magnetic enzyme immunoassay technique to detect standard solution for Shigella （ATCC 25931 ）, result showed that correlation coefficient between relative light intensity detected by instrument and bacteria concentration detec- ted by culture counting method was 0.981 1. Moreover, relation curve between relative light intensity and Shigella concentration was drawn. [ Conclusion] The method had a high detection speed and accuracy, and could be used for the rapid detection of pathogen in food and environment.

Rapid bioluminescence assay for monitoring rat CES1 activity and its alteration by traditional Chinese medicines

In traditional Chinese medicine herbs(TCM),including Radix Salviae Miltiorrhizae(Danshen),Radix Puerariae Lobatae(Gegen),Radix Angelicae Sinensis(Danggui),and Rhizoma Chuanxiong(Chuanxiong)are widely used for the prevention and treatment of cardiovascular diseases and also often co-administered with Western drugs as a part of integrative medicine practice.Carboxylesterase 1(CES1)plays a pivotal role in the metabolisms of pro-drugs,Since(S)-2-(2-(6-dimethylamino)-benzothiazole)-4,5-dihydrothiazole-4-carboxylate(NLMe)has recently been identified by us as a selective CES1 bioluminescent sensor,we developed a rapid method using this substrate for the direct measurement of CES1 activity in rats.This bioluminescence assay was applied to determine CES1 activity in rat tissues after a two-week oral administration of each of the four herbs noted above.The results demonstrated the presence of CES1 enzyme in rat blood and all tested tissues with much higher enzyme activity in the blood,liver,kidney and heart than that in the small intestine,spleen,lung,pancreas,brain and stomach.In addition,the four herbs showed tissue-specific effects on rat CES1 expression.Based on the CES1 biodistribution and its changes after treatment in rats,the possibility that Danshen,Gegen and Danggui might alter CES1 activities in human blood and kidney should be considered.In summary,a selective and sensitive bioluminescence assay was developed to rapidly evaluate CES1 activity and the effects of orally administered TCMs in rats.

Source reconstruction for bioluminescence tomography via L_(1/2)regularization

Bioluminescence tomography(BLT)is an important noninvasive optical molecular imaging modality in preclinical research.To improve the image quality,reconstruction algorithms have to deal with the inherent ill-posedness of BLT inverse problem.The sparse characteristic of bioluminescent sources in spatial distribution has been widely explored in BLT and many L1-regularized methods have been investigated due to the sparsity-inducing properties of L1 norm.In this paper,we present a reconstruction method based on L_(1/2) regularization to enhance sparsity of BLT solution and solve the nonconvex L_(1/2) norm problem by converting it to a series of weighted L1 homotopy minimization problems with iteratively updated weights.To assess the performance of the proposed reconstruction algorithm,simulations on a heterogeneous mouse model are designed to compare it with three representative sparse reconstruction algorithms,including the weighted interior-point,L1 homotopy,and the Stagewise Orthogonal Matching Pursuit algorithm.Simulation results show that the proposed method yield stable reconstruction results under different noise levels.Quantitative comparison results demonstrate that the proposed algorithm outperforms the competitor algorithms in location accuracy,multiple-source resolving and image quality.

In vivo bioluminescence imaging of hyperglycemia exacerbating stem cells on choroidal neovascularization in mice

AIM： To investigate the influence of hyperglycemia on the severity of choroidal neovascularization（CNV）,especially the involvement of bone marrow-derived cells（BMCs） and underlying mechanisms.·METHODS： BMCs from firefly luciferase（Fluc）/green fluorescent protein（GFP） double transgenic mice were transplanted into C57BL/6J wide-type mice. The recipient mice were injected intraperitoneally with streptozotocin（STZ） daily for 5 consecutive days to induce diabetes mellitus（DM）, followed by CNV laser photocoagulation.The BMCs recruitment in CNV exposed to hyperglycemia was firstly examined in Fluc/GFP chimeric mice by in vivo optical bioluminescence imaging（BLI） and in vitro Fluc assays. The CNV severity was evaluated by H＆E staining and choroidal flatmount. The expression of vascular endothelial growth factor（VEGF） and stromal cell derived factor-1（SDF-1） was detected by Western blot.·RESULTS： BLI showed that the BMCs exerted dynamic effects in CNV model in Fluc/GFP chimeric mice exposed to hyperglycemia. The signal intensity of transplanted Fluc＋GFP＋BMCs in the DM chimeric mice was significantly higher than that in the control chimeric mice with CNV induction at days 5, 7, 14 and 21（121861.67 ±9948.81 vs 144998.33 ±13787.13 photons/second/cm2/sr for control and DM mice, P5d〈0.05; 178791.67±30350.8 vs240166.67 ±22605.3, P7d〈0.05; 124176.67 ±16253.52 vs196376.67 ±18556.79, P14d〈0.05; 97951.60 ±10343.09 vs119510.00 ±14383.76, P21d〈0.05）, which was consistent with in vitro Fluc assay at day 7 [relative light units of Fluc（RLU1）], 215.00±52.05 vs 707.33±88.65, P 〈0.05; RLU1/relative light units of renilla luciferase（RLU2）, 0.90 ±0.17 vs 1.83 ±0.17, P 〈0.05]. The CNVs in the DM mice were wider than those in the control group at days 5, 7, 14 and21（147.83±17.36 vs 220.33±20.17 μm, P5d〈0.05; 212.17 ±24.63 vs 326.83 ±19.49, P7d〈0.05; 163.17 ±18.24 vs265.17 ±20.55, P14d〈0.05; 132.00 ±10.88 vs 205.33 ±12.98,P21d〈0.05）. The average area of CNV in the DM group was larger at 7d（20688.67±3644.96 vs 32218.00±4132.69 μm2,P 〈0.05）. The expression of VEGF and SDF-1 was enhanced in the DM mice.·CONCLUSION： Hyperglycemia promots the vasculo-genesis of CNV, especially the contribution of BMCs,which might be triggered by VEGF and SDF-1 production.

The Effects of Trehalose on the Bioluminescence and Pigmentation of the Phase I Variant of Photorhabdus luminescens

Photorhabdus luminescens is a Gram-negative, bioluminescent, pigment producing enteric bacterium, which is pathogenic to insects and has the capability to undergo phase variation. The phase I variant of P. luminescens exists as a mutualistic symbiont where it plays a critical role in the life-cycle of the soil-dwelling nematode, Heterorhabditis bacteriophora. Both the bacterium and the nematode receive their nutritional requirements from the bioconversion of the insect host which is rich in many macromolecules such as the disaccharide, trehalose. Trehalose is a non-reducing disaccharide of glucose that is formed by an a-1,1-glycosidic bond and is associated with the physiology of many bacteria, insects and nematodes. Trehalose has been shown to be the most abundant storage sugar found within insect hemolymph （1%-2%）. The physicochemical properties of trehalose allow this carbohydrate to act as a stress protectant where it has been implicated with thermal stress, dehydration, and osmotic protection of many microorganisms. Due to these properties, trehalose may allow culture stability of the phase I variant in vitro and in vivo. Traits of the phase I variant that were studied in this work include bioluminescence and the production of the red anthroquinone-derived pigment. The carbohydrates that were utilized in this study were glucose and trehalose; where shake flask cultures of the phase I variant were cultured at room temperature for up to six days in carbohydrate supplemented basal media with increasing carbohydrate concentrations of 0. 1%, 0.5% and 1.0% （v/v）. Relative luminosity, pigmentation and pH were graphed as a function of time, carbohydrate used, and carbohydrate concentration. Data obtained from this study suggests that the supplementation of 1.0% trehalose, when culturing the phase I variant ofP. luminescens, can maintain bioluminosity and pigmentation over extended periods of time （five days） as compared to basal media and basal media supplemented with 1.0% glucose.

Application value of ATP based bioluminescence tumor chemosensitivity assay in the chemotherapy for hydrothorax caused by non-small cell lung cancer

Objective： The aim of the study was to investigate the clinical value and application of ATP based bioluminescence tumor chemosensitivity assay （ATP-TCA） in the chemotherapy for hydrothorax caused by non-small cell lung cancer （NSCLC）. Methods： Hydrothorax specimens from 120 NSCLC patients were analyzed by ATP-TCA and the most sensitive chemotherapeutic drugs were used in NSCLC patients （treatment group）. At the same time, 56 NSCLC patients with hydrethorax were admitted in our Hospital （Department of Oncology, The No. 2 People＇s Hospital of Yibin, China） and given chemotherapy without guidance of the ATP-TCA （control group）. Before the third chemotherapeutic cycle, clinical outcomes were analyzed in the two groups. Results： Effective rate of hydrothorax in treatment group was 67%, while 46% in control group （P 〈 0.05）. In refractory hydrothorax patients, they were 69% and 40% （P 〈 0.05）, respectively.In vitro results correlated well with clinical outcomes （P 〈 0.01）. Conclusion： Effective rate of chemotherapy for hydrothorax in NSCLC is higher in treatment group than that in control group. ATP-TCA is especially helpful for refractory hydrothorax.

Three-Dimentional Surface Light Irradiance Reconstruction in Bioluminescence Tomography

Reconstruction of 3D surface irradiance distribution using multiple views captured by charged coupled device(CCD)camera is the basis of solving the light source in bioluminescence tomography(BLT).A simple and convenient mapping technique based on the pin-hole imaging model and Lambert′s cosine law was presented to establish the relationship between gray levels and irradiance intensities.Compared with previous integrating sphere camera calibration used in BLT,the proposed method can effectively avoid heavy burden of simulation experiment to obtain the corresponding relationship of gray levels and irradiance intensities.The accuracy and feasibility of the proposed method are validated with no more than 1mm location error by different types of phantom experiments.The mapping approach is also applicable to other noncontact optical imaging system.

Fast in vivo bioluminescence tomography using a novel pure optical imaging technique

Bioluminescence tomography(BLT)is a novel opt ical molecular imaging technique that advanced the conventional planar bioluminescence imaging(BLI)into a quantifiable three-dimensional(3D)approach in preclinical living animal studies in oncology.In order to solve the inverse problem and reconstruct tumor lesions inside animal body accurately,the prior structural information is com-monly obtained from X ray computed tomography(CT).This strategy requires a complicated hybrid imaging system,extensive post imaging analysis and involvement of ionizing radiation.Moreover,the overall robustness highly depends on the fusion accuracy between the optical and structural information.Here,we present a pure optical bioluminescence tomographic(POBT)system and a novel BLT workfow based on multi-view projection acquisition and 3D surface reconstruction.This met hod can reconstruct the 3D surface of an imaging subject based on a sparse set of planar white-light and bioluminescent images,so that the prior structural information can be offered for 3D tumor lesion reconstruction without the involvement of CT.The performance of this novel technique was evaluated through the comparison with a conventional dual-modality tomo-graphic(DMT)system and a commercialized optical imaging system(IVIS Spectrum)using three breast cancer xenografts.The results revealed that the new technique offered comparable in vivo tomographic accuracy with the DMT system(P>0.05)in much shorter data analysis time.It also offered significantly better accuracy comparing with the IVIS system(P<0.04)without sacrificing too much time.

Influence of the temperature at the Black Sea ctenophores-aliens bioluminescence characteristics

Successful invasion of Mnemiopsis leidyi A. Agassiz, 1865 and Beroe ovata Mayer, 1912 into the Black Sea and their important role in this region pelagic ecosystem is stipulated mainly by the considerable eurythermy of these species. Many ecological-physiological characteristics of ctenophores—aliens are studied quite well. However, bioluminescence, one of the most important elements of the ctenophores ecology and the bioluminescence reaction temperature optimum for these individuals under different environment temperatures were not studied sufficiently. Therefore our researches in this scientific field are significant and conceptually novel for ctenophores ecology study. Experimental investigations were carried out in the period of 2008-2009 in the IBSS. Uni-sized (40 mm) ctenophores were collected in the Sevastopol coastal zone and divided in several groups, contained under different temperatures: from 10°C ± 1°C to 30°C ± 1°C. Ctenophore bioluminescence was investigated under chemical and mechanical stimulation. M. leidyi light emission maximal amplitude (1432.94 ± 71.64 × 108 quantum·s–1·cm–2) with duration of 3.54 ± 0.15 s is fixed under the temperature of 26°C ± 1°C. Temperature increase up to 30°C ± 1°C led to the 4 times decrease of the bioluminescence intensity. Under temperature decrease up to 10°C ± 1°C this parameter decreased 20 times (p B. ovata achieved maximal values under the temperature of 22°C ± 1°C (1150.12 ± 57.51 × 108 quantum·s–1·cm–2) with duration of 3.03 ± 0.15 s. The luminescence intensity decreased under the temperature increase to 30°C ± 1°C more than 20 times. Temperature decrease to the values of 10°C ± 1°C impacted decreasing the amplitude of bioluminescence up to the minimal –4.92 ± 0.22 × 108 quantum·s–1·cm–2. The data obtained testify that characteristics of the ctenophores bioluminescence can be conditioned not only by the modification the environment temperature but by the variability of their physiological condition.

Variability of the bioluminescence characteristics of the Black Sea ctenophores-aliens in connection with different conditions of nutrition

Many ecological-physiological characteristics of ctenophores-aliensMnemiopsis leidyi A. Agassiz, 1865 and Beroe ovata Mayer, 1912 are studied quite well because they play a very important ecological role in the Black Sea ecosystem. However, bioluminescence, one of the most important elements of the ctenophores ecology and its connection with feeding regime were not studied sufficiently. Experiments have shown that characteristics of the ctenophores bioluminescence differed considerably in dependence of food supplies. Thus, amplitude and light-emitting energy of the fed ctenophores B. ovata are maximal, 3 times more than analogical indices of the just-caught individuals and 4 times more than ones of starving individuals. More prolonged flash signal (to 3.5 s), which exceeds light-emitting duration of the starving individuals twice, can be registered from the fed ctenophores. Investigation of the M. leidyi bioluminescence has shown that amplitude and light-emitting energy of the just-caught ctenophores were two times more than those of the starving individuals. At the same time, light-emitting amplitude of the fed individuals is 6.5 times and light-emitting energy is 3 - 4 times higher than that of the just-caught ctenophores. The light-emitting duration of the starving and justcaught organisms is practically the same. The most prolonged signal is registered from the fed ctenophores—up to 2.8 s. The data obtained testify that characteristics of the ctenophores bioluminescence can be conditioned not only by nutritional value but by the composition of the food as well.

Some Theoretical and Applied Aspects of the Oceanic Bioluminescence Registration from Space

The analysis of the surface bioluminescence in the World Ocean registration problems for the purposes of its regions ecological express-monitoring and evaluation of the plankton algae spatial distribution at night time by the existing space systems has been proposed. The connection of the plankton community characteristics with a bioluminescent potential (BP) in the euphotic layer and a possibility to evaluate BP according to bioluminescence intensity in the near-surface layer of 0 - 10 m has been demonstrated. It has been shown that with complete correspondence with the vertical structure in the plankton community at the dark time bioluminescence intensity in 0 - 10 m layer exceeds the same in 60 - 70 m layer for one and a half orders and practically determines BP in 0 - 100 m layer. Peculiarities of the plankton organisms light emission, important for the oceanic bioluminescence registration with the space means of observation are under discussion. Equation for calculation of the measured by the space device sea luminescence level and volume of the bioluminescence intensity in the surface layer of theWorldOcean, which can be registered due to modern technical means of the space systems has been corrected. The conclusion has been done that on the base of the space data about spatial and temporal phytoplankton distribution and bioluminescence “in situ” measurements it is possible to create regional algorithms for transition from numerical estimations of the phytoplankton (chlorophyll “a”) to the day time bioluminescent potential and solution of inverse tasks at night time.

Bioluminescence and Chitinase Production during Chitin Fermentation by Vibrio harveyi

Vibrio harveyi, like other luminescent bacteria, is capable of producing extracellular chitinases. Microbial chitinases are utilized to depolymerize chitin into chitooligosaccharides and N-acetylglucosamine for the acquisition of carbon and possibly nitrogen, needed for survival. For many luminous marine bacteria （Vibrio spp.）, quorum-sensing is highly speculated to be responsible for bioluminescence; however, in terrestrial species （Photorhabdus spp.） luminosity seems to be controlled through unknown mechanism of phase variation. In the present work, the correlation between bacterial luminosity and chitinase production of F. harveyi was studied. The utilization of bioluminescence could prove to be an easier and more convenient method to monitor chitin fermentations that employ luminous bacteria. Results from the fermentation study indicate that luminosity of F. harveyi inversely correlates with chitinase production. In other words, during chitin fermentation, chitinase production was seen to increase while luminosity decreased with respect to growth and growth conditions. Furthermore, the results also suggest that V. harveyi may utilize an alternate mechanism that can counter quorum-sensing mechanisms to ensure bacterial survival under deteriorating growth conditions. The inverse relationship observed in this study may lead to a basic understanding of monitoring and studying chitin fermentations and anti-quorum-sensing/phase variation mechanisms exhibited by luminous bacteria.

Differential Regulation of Proteins and a Possible Role for Manganese Superoxide Dismutase in Bioluminescence of Panellus stipticus Revealed by Suppression Subtractive Hybridization

Suppression subtractive hybridization (SSH) was employed to investigate bioluminescence in Panellus stipticus (Bull.) P. Karst. by detecting proteins differentially expressed in bioluminescent and luminescent strains. Comparisons of luminescent and non-luminescent monokaryon cultures of North American strains revealed differences in transcript levels of proteins responsible for post-translational modification (PTM) of enzymes. A similar comparison of a luminescent strain of P. stipticus from North America with a non-luminescent European strain revealed the presence of extracellular manganese superoxide dismutase (MnSOD) in the luminescent form, in addition to proteins involved in PTM. The application of MnSOD-specific inhibitors to luminescent mycelium resulted in the rapid loss of luminescence. The relevance to luminescence of proteins involved in PTM is discussed, together with a possible role for MnSOD that considers the potential for SODs to form stable complexes with catechols revealed in previously published research. In light of the recent discovery that hispidine may be the precursor of fungal luciferin, we consider a hypothetical mechanism for fungal luminescence in which the ο-hydroquinone moiety of a hispidine derivative ligates with the extracellular form of MnSOD producing a semiquinone-radical complex, with the resultant semiquinonato complex potentially reacting with molecular oxygen or other reactive oxygen species to produce sufficiently excited intermediates to emit light on relaxation.

Luminous fishes:Endocrine and neuronal regulation of bioluminescence

Bioluminescence is the ability of organisms to emit their own light which can be self-produced in light organs by photogenic cells(photocytes)or symbiotically by bearing cultures of luminous bacteria.Luminous animals are mainly comprised in the ocean,nocturnal coastal shallow-water species or,as the majority of species,inhabiting deep-sea where dim light conditions are optimal for communication by luminous signals.Due to difficult access to species,fisheries of bioluminescent fishes worldwide are rare,mostly for traditional cuisine and fishing bait.Fishes are the only bioluminescent vertebrates and thus their light organs present an astonishing level of complexity for light modulation to adequate light signals for specific ecological roles.Fishes control the output of light intensity using different mechanisms which can be categorized as being under control of hormonal or neuronal pathways.Usually,neural control allows fast and direct response to the photocytes while endocrinal systems allow to control the light produced by bacteria,via manipulation of individual melanophores.In some species,mechanical movement of melanophore-filled tissues by muscular action is under neuronal or hormonal control.Sexual dimorphism of light organs,driven by hormonal regulation during ontogeny,takes a crucial role in communication and reproductive success of these species.

Is deep brain imaging on the brink of transformation with a bioluminescence molecule?

Cephalofurimazine(CFz),when paired with Antares luciferase,shows superior blood-brain barrier permeability and enhanced imaging depth and clarity for deep brain imaging.This bioluminescence provides a less invasive method for real-time monitoring of deep brain activity,with the potential to advance targeted therapies and deepen our understanding of brain functions.Further molecular engineering and localized delivery can reduce the potential toxicity of CFz and enhance its efficacy for clinical deep brain imaging.

Optimizing a high-sensitivity NanoLuc-based bioluminescence system for in vivo evaluation of antimicrobial treatment

Focal and systemic infections are serious threats to human health.Preclinical models enable the development of new drugs and therapeutic regimens.In vivo,animal bioluminescence(BL)imaging has been used with bacterial reporter strains to evaluate antimicrobial treatment effects.However,high-sensitivity bioluminescent systems are required because of the limited tissue penetration and low brightness of the BL signals of existing approaches.Here,we report that NanoLuc(Nluc)showed better performance than LuxCDABE in bacteria.However,the retention rate of plasmid constructs in bacteria was low.To construct stable Staphylococcus aureus reporter strains,a partner protein enolase(Eno)was identified by screening of S.aureus strain USA300 for fusion expression of Nluc-based luciferases,including Nluc,Teluc,and Antares2.Different substrates,such as hydrofurimazine(HFZ),furimazine(FUR),and diphenylterazine(DTZ),were used to optimize a stable reporter strain/substrate pair for BL imaging.S.aureus USA300/Eno-Antares2/HFZ produced the highest number of photons of orange-red light in vitro and enabled sensitive BL tracking of S.aureus in vivo,with sensitivities of approximately 10 CFU from mouse skin and 750 CFU from mouse kidneys.USA300/Eno-Antares2/HFZ was a powerful combination based on the longitudinal evaluation of the therapeutic efficacy of antibiotics.The optimized S.aureus Eno-Antares2/HFZ pair provides a technological advancement for the in vivo evaluation of antimicrobial treatment.

Bio-screening and quantification of methyl paraben in vinegar and coconut juice separated by HPTLC

As a widely used food preservative,methyl paraben was experimentally evidenced with serious hormonelike adverse effects.Herein,a high performance thin-layer chromatography platformed bioluminescent bioautography and image analysis for the selective quantification and confirmation of methyl paraben was proposed and validated in vinegar and coconut juice.First,the detectability of the bioautography to the analyte on different layer materials was estimated,revealing that normal silica gel was the best choice.After that,the liquid of sample extract and working solution were separated to overcome the background noises due to co-extracted matrices.The separation result was then coupled to the optimized bioautography,enabling instant and straightforward screening of the targeted conpound.For accurate quantification,bioluninescent inhibition pattern caused by the analyte was processed by image analysis,giving useful sensitivity(LOD>16 mg/kg),precision(RSD<10.1%)and accuracy(spike-recovery rate 76.9%-112.2%).Finally,the suspected result was confirmed by determining its MS fingerprint,further strengthening the reliability of screening.

Optical molecular imaging in cancer research:current impact and future prospect

Cancer has long been amajor threat to human health.Recent advancements inmolecular imaging have revolutionized cancer research by enabling early and precise disease localization,essential for effective management.In particular,optical molecular imaging is an invaluable cancer detection tool in preoperative planning,intraoperative guidance,and postoperative monitoring owing to its noninvasive nature,rapid turnover,safety,and ease of use.The tumor microenvironment and cells within it express distinct biomarkers.Optical imaging technology leverages these markers to differentiate tumor tissues from surrounding tissues and capture real-time images with high resolution.Nevertheless,a robust understanding of these cancer-relatedmolecules and their dynamic changes is crucial for effectivelymanaging cancer.Recent advancements in opticalmolecular imaging technologies offer novel approaches for cancer investigation in research and practice.This review investigates themodern opticalmolecular imaging techniques employed in both preclinical and clinical research,including bioluminescence,fluorescence,chemiluminescence,photoacoustic imaging,and Raman spectroscopy.We explore the current paradigm of optical molecular imaging modalities,their current status in preclinical cancer research and clinical applications,and future perspectives in the fields of cancer research and treatment.

Application of stimulating agents on the immobilized bioluminescence strain Pseudomonas putida mt-2 KG1206,preserved by deep-freezing,for the convenient biomonitoring

This study was conducted to develop methods for the application of an immobilized bioluminescence strain （KG1206）, preserved by deep-freezing （DF）, for the monitoring of contaminated environments. The immobilized cells, preserved by DF, required approximately 2 hr for reconstitution of their activity. A large reduction in bioluminescence was observed due to the DF process; 0.07-0.58 times that of the non deep-frozen （NDF） immobilized strain. The decreased bioluminescence activity induced by the DF process was enhanced by the stimulants, sodium lactate （SL） and KNO3. However, regardless of the inducer chemical tested, the immobilized strain modified with KNO3 consistently produced greater bioluminescence than that treated with SL, in the range of 3.0-10.7 （avg. 6.7± 3.69） and 1.2-4.2 （avg. 2.4 ± 1.47） times that of control, respectively. All KNO3 treatments of contaminated groundwater samples also resulted in an increase in bioluminescence activity, but the rate of stimulation varied for each sample. Also, no strong linear correlation was observed between the bioluminescence and the total concentration of an inducer, which may related to the complex characteristics of the environmental samples. Overall, the results demonstrated the ability of immobilized genetically engineered bacteria, preserved by DF, to measure a specific group of environmental contaminants using a stimulating agent （KNO3）, suggesting the potential for its preliminary application in a field-ready bioassay.