Background and Objective Chronic and infected leg ulcers (LUs) are painful, debilitating, resistant to antibiotics, immensely reducing patient's quality of life. The purpose of the new clinical research project was to demonstrate the the efficacy of photodynamic therapy (PDT) as an alternative to antibiotic therapy for the treatment of infected chronic LUs.
Patients and Methods PDT using irradiation at fluency 80 J / cm2 and 20% 5-aminolevulinic acid topically applied for 4 hrs, was performed during 10 days of hospitalization in 10 patients both sexes aged 40-85, with chronic leg ulcers, who had not responded well to conventional treatment. This was a randomized, placebo-controlled trial (local Octenidine dihydrochloride). The numerical scale of pain was used. Treatments were carried out at 3-week intervals.
Result During 8 months follow up, complete remission (CR) was obtained in 4 patients (40%), partial response (>50% reduction in ulcer diameter), in 3 patients (30%), no response in 1 patients (10%). In two patients (20%) was observed deterioration of the local condition, with swelling, erythema and inflammation.
Conclusion PDT allowed healing of chronic leg ulcers through the antimicrobial effect, modulation of inflammatory process and improving the oxygenation of ulcer tissue with a reduction in healing time and improving scarring results with the reduction of pathological bacterial culture. Further studies on the larger group of patients are needed to confirm the benefit role of PDT in the treatment of LUs.
The aim of this study was to determine the immune effect of PDT with 5-aminolevulinic acid (ALA-PDT) used in sublethal doses on the secretion of growth factors: VEGF, GM-CSF, G-CSF, FGF, Interleukin 6,8 and 10 and S100 protein secretion by experimental models of residual colon cancer cells in vitro.
Background and objective The search for new diagnostic and therapeutic procedures is an essential task in contemporary oncology. The purpose of our study was the evaluation of the practical usefulness of endoscopic autofluorescence assessment (AFE) using the Onco-LIFE system, and the estimation of the correlation between the histopathological evaluation with the degree of lesions’ Numerical Color Value (NCV index) and the method’s sensitivity and specificity valuation.
Material 67 patients were analyzed and had previously a tumor of the gastrointestinal tract which appeared malignant, but without histopathological confirmation. We measured NCV, estimated the correlation of the clinical diagnosis based on histopathological evaluation with the degree of NCV index from gastrointestinal lesions, and calculated the sensitivity and specificity of this method.
Results In the group of 67 patients, we found 44 cases of primary or secondary cancers and 7 cases of non-epithelial malignancies. In this group (51 patients) we identified 13 colorectal cancers and 38 upper gastrointestinal cancers. Based on the NCV index at NCV>1.0, we can show that the sensitivity for malignant neoplastic lesions was 100% and the specificity was 73%, while for NCV>1.5, the sensitivity for malignant neoplastic lesions was 86% and the specificity 100%.
Conclusion AFE using the Onco-LIFE system is a helpful tool to perform targeted biopsies at the outset. A significant correlation was found between lesions’ NCV index and the grade of dysplasia or tumor malignancy. AFE sensitivity and specificity is higher than WLE. Further studies are needed, especially performed by expert endoscopists.
Aim of the study was to determine the immune effect of PDT with hypericin (HY-PDT) used in sublethal doses on the secretion of Interleukin 8 and 10 by experimental models of residual colon cancer cells in vitro.
Results HY-PDT amplified the secretion of IL-8 by SW620 cell line, but the decrease of IL-8 secreted by the SW480 cell line. The increase in secretion of IL-10 was noticed in the SW480 cell line, but the changes of secretion IL-10 by SW 620 was not noted. SW480 cell line without PDT secreted higher levels of IL-8 and IL-10 than SW620 cells.
Based on these research findings it could be told, that PDT both eliminates and control primary tumors using cytotoxic effect while HY-PDT at lower doses can modulate function of tumor microenvironment by releasing interleukins depended on metastatic activity of tumor cells.
Background. This study assessed the efficacy of autofluorescence endoscopy (AFE) using the Onco-LIFE system and numerical color value (NCV) estimation in comparison to white light endoscopy (WLE) in endoscopic surveillance for identification of early dysplasia in Barrett’s esophagus (BE). In the case of dysplasia photodynamic treatment (PDT) was carried out.
Methods. AFE, performed simultaneously during WLE, with biopsy was performed in 24 patients with BE. None of these patients had any obvious mucosal abnormalities in WLE. A total of 376 biopsies were taken, include 325 randomly collected according to Seattle Protocol (SP) and 51 additional biopsies, taken from the sites with pathological AF and NCV, and then compared to histological examinations. The PDT was performed using 5-aminolevulinic acid (ALA) and DIOMED 630nm light source.
Results. In the case of 248 biopsies taken from sites with NCV below 1.0, two cases of unspecified dysplasia were recognized; in 14 biopsies with NCV above 2.0 in all cases the various grades of dysplasia were documented. Dysplasia was found in 42% of AFE+NCV- guided biopsy specimens, and in 7.1% of WLE-guided biopsy specimens. AFE+NCV detected high-grade dysplasia in 7 patients, 6 more than according to SP in WLE. In the group patients with dysplasia, PDT was successfully carried out.
Conclusion: The results indicate that AFE+NCV using the Onco-LIFE system leads to improved BE lesion visualization for targeted biopsy with accurate histologic correlation compared to WLE and SP guided biopsy, and can serve to minimize additional biopsies. PDT is an effective therapeutic method in BE.
Solitary rectal ulcer syndrome (SRUS) is a chronic disease of the rectum. Although SRUS is a benign condition there are
studies which suggest that chronic ischaemia which occurs in the SRUS may lead to "transitional mucosa" that is similar
to that adjacent to colorectal carcinomas and adenomas and may lead to colorectal dysplasia and carcinoma development.
The exclusion of primary or metastatic malignancy is the most important aim in the differential diagnosis of SRUS. In our
study we assess the possibilities of autofluorescence colonoscopy (AFC) in diagnosis and management of SRUS.
We performed white light colonoscopy first. The tissue samples were taken for pathological examination. When SRUS
was histopathologically confirmed AFC was performed by means of Xillix OncoLIFE. During AFC numerical colour
value (NCV) of autofluorescence of SRUS lesions was noted.
During 1946 colonoscopies eight persons were diagnosed as having solitary rectal ulcer syndrome. We did not observe
autofluorescence increase in case of polipoid and flat ulcer lesions (NCV 0,39-0,67; mean 0,525) and little increase of
autofluorescence in case of erythema lesion (NCV- 0,94).
SRUS is a rare disorder of the rectum but it causes differential diagnosis problems. The most common reason for
incorrect diagnosis are inadequate tissue specimens. AFC allows to reveal subtle areas within the lesions of more intense
autofluorescence and localizes the potential cancer-transformating dysplasia. In this way the most representative area
with highest risk of pre- or cancerous changes, for biopsy specimen is indicated.
Background: Fluorescence diagnostics uses the ability of tissues to fluoresce after exposition to a specific wavelength of
light. The change in fluorescence between normal and progression to cancer allows to see early cancer and precancerous
lesions often missed by white light.
Aim: To improve by computer image processing the sensitivity of fluorescence images obtained during examination of
skin, oral cavity, vulva and cervix lesions, during endoscopy, cystoscopy and bronchoscopy using Xillix ONCOLIFE.
Methods: Function of image f(x,y):R2 → R3 was transformed from original color space RGB to space in which vector of
46 values refers to every point labeled by defined xy-coordinates- f(x,y):R2 → R46. By means of Fisher discriminator
vector of attributes of concrete point analalyzed in the image was reduced according to two defined classes defined as
pathologic areas (foreground) and healthy areas (background). As a result the highest four fisher's coefficients allowing
the greatest separation between points of pathologic (foreground) and healthy (background) areas were chosen. In this
way new function f(x,y):R2 → R4 was created in which point x,y corresponds with vector Y, H, a*, c2.
In the second step using Gaussian Mixtures and Expectation-Maximisation appropriate classificator was constructed.
This classificator enables determination of probability that the selected pixel of analyzed image is a pathologically
changed point (foreground) or healthy one (background). Obtained map of probability distribution was presented by
means of pseudocolors.
Results: Image processing techniques improve the sensitivity, quality and sharpness of original fluorescence images.
Conclusion: Computer image processing enables better visualization of suspected areas examined by means of
fluorescence diagnostics.
The aim of the experiment was to estimate the effect of repeated simultaneous exposures to infrared laser radiation and μT variable magnetic field used in magnetostimulation on pain perception in rats, as well as the involvement of endogenous opioid system in the mechanism of this effect. In experimental group clean-shaven scull of male Wistar rats placed individually in a specially designed plastic chamber were simultaneously exposed to infrared laser radiation
(wavelength - 855 nm, mean power - 4,1 mW, energy density - 30 J/cm2) and variable magnetic field of saw-like shape
of impulse, at a frequency of basic impulse 180-195 Hz and mean induction value of 120 μT generated by magneto-laser
applicator of device for magnetostimulation Viofor JPS (Med & Life, Poland) 12 minutes daily for 2 periods of 5 consecutive days, with 2 days-lasting break between them, while control animals were sham-exposed. The pain perception was determined by means of "hot plate" test on the basis of calculated analgesic index. As a result of repeated exposures a significant increase in analgesic index persisting also till 14th day after the end of a cycle of exposures was observed. This analgesic effect was inhibited by prior i.p. injection of opioid antagonist - Naloxone.
Introduction. Life induced fluorescence (LIFE) diagnostics can be used as an imaging system of
precancerous and neoplastic lesions of the oral mucosa. Neoplastic lesions are visible in pseudo colours,
healthy tissue in green colour and abnormal tissue in red colour. All the observed colours present
different intensity. Colour intensity is relevant to the grade of dysplasia, carcinoma progress and is called
Numerological Value of Color Index (NCV). The aim of our study was to find correlation between
autofluorescence diagnostics combined with NCV assessment and histopathological findings of taken
specimen biopsies. Patients and methods. 10 patients participated in our study. Lesions affected a variety of
intraoral sites. The most common location was: buccal, gingival and mandibular mucosa. Patients were
examined using Life Induced Fluorescence diagnosis (400 - 750 nm wavelength) with Numerological
Value of Color index (NCV) using Onco LIFE system. Afterwards the specimen biopsies from the lesions
were taken and histopathological examination was performed. Results. Different NCV and dependence of
NCV on the histopathological findings were observed. Conclusion. Diagnostic procedures with the
application of white-light imaging with LIFE imaging is not only a significantly faster method and a
better diagnostic tool of preneoplastic and neoplastic lesions, but there exist also correlations between
measured NCV and histopathological diagnosis. The farther investigations are necessary in order to prove
these preliminary findings.
Introduction. The whole mechanism of the cellular level of tumor destruction by photodynamic therapy (PDT) is still
unknown. Despite necrotic and apoptotic ways of cell death, there is a variety of events leading to and magnifying the
inactivation of tumor cells. Material and methods. J-774A.1 were incubated with δ-aminolevulinic acid (ALA) at different concentrations (125, 250, 500, 1000 μM) and then irradiated with VIS (400 - 750 nm) at the dose of 5,10 and 30 J/cm2 delivered from the incoherent light source. The effects of the application of ALA-PDT were evaluated on the basis of cell viability,
nitric oxide (NO), tumor necrosis factor α- (TNF-α) and interleukin-1β (IL-1β) produced by the J-774A.1 cells. Results. The cell viability (assessed using MTT test) was comparable with control group at 5,10 and 30 J/cm2. At these doses of
energy using different concentrations of ALA we have observed that at the higher energy doses, the greater increase of TNF-α release, lowering of the level of IL-1β production and decrease of NO release were observed. There was also observed the dependence of the secretional activity of the cells on the ALA concentrations. Conclusion. The cell viability and production of cytokines depended on ALA concentrations and energy doses of the light. The higher some cytokines' release after PDT could be an additional factor for the complete eradication of tumor.
Photodynamic therapy (PDT) is one of a novel modalities which has recently been exploited to eradicate various microorganisms. In our study we have evaluated bactericidal efficacy of PDT in the presence of 5-δ aminolaevulinic acid (ALA). Porphyromonas gingivalis were incubated with increasing concentration of ALA and subsequently irradiated by progressive light doses. Complete killing effect was obtained for bacteria irradiated with 25J/cm2 in ALA solution final concentration of 1mM, 5mM, 10mM. Statistical analysis has revealed ALA concentration to be a major factor responsible for eradication of bacteria. The latter may be attributable to the known ALA dark toxicity.
The results of application of photodynamic diagnostics (PDD) and photodynamic therapy (PDT) for diagnosis and treatment of premalignant lesions and malignant tumours of different tissues were presented. In the period 1998-2000 a total of 314 patients were diagnosed with use of different methods of PDD and a total of 78 patients were treated with use of PDD. On the basis of obtained result we consider PDD and PDT to be sensitive, efficient and non-invasive methods of diagnostics and therapy of malignant lesions.
110 patients were subjected to colonscopy under white light and autofluorescence. An adapted Xillix LIFE system and xenon light source with interference filter were used in the autofluorescence diagnostics. Both techniques revealed 67 small lesions, including 45 neoplastic ones and 22 others in the examined patients. The autofluorescence examination revealed 43 neoplastic changes, while conventional white-light colonscopy allowed for localization of 39 neoplastic lesions. Autofluorescent imaging of large intestine mucosa is more sensitive in the detection of small neoplastic changes than traditional colonscopy.
The efficiency of autofluorescence diagnosis within urinary bladder was analyzed in the study. We examined two groups of patients: the first one consisting of 22 patients suspected to have bladder cancer and the second one consisting of 45 patients who have undergone transurethral electro resection due to urinary bladder neoplasms. Our goal was to detect cancerous tissue invisible in white-light examination. In the first group sensitivity was 100 percent and specificity was 69.23 percent. In the second group sensitivity was 96 percent and specificity was 80 percent. We also report in the study treatment efficiency of PDT in 12 patients with superficial bladder cancer. In our procedure two hours after the instillation of bladder with ALA solution, the lesion was irradiated by laser light. In 9 out of 12 treated patients regression of bladder tumor was obtained, while in 3 cases a progression of neoplasmatic process was observed.
In this paper the authors present their own experience in lasertherapy of motional system overloading syndromes in 522 patients, in which laser radiation was used as monotherapy. IR semiconductor lasers and He-Ne laser were used. Each irradiation procedure was performed in 2 stages: 1) the scan of painful region; 2) irradiation of trigger points. As a result of 10-25 irradiations the improvement was obtained in 61-87 percent of patients and it was significantly better as compared to the control group in which sham-irradiation was made. On the basis of obtained results one may conclude that lasertherapy is a very useful, assisting method in treatment of motional system overloading syndromes. The therapeutical effects of the method are related to analgesic and antiphlogistic action of laser radiation.
The aim of the study was to confirm the analgesic effect of low-power laser radiation with a tail-immersion test and check if nitric oxide is involved in laser radiation-induced analgesia in rats. The experiment was performed on male Wistar rats. On the day of experiment the scull of rats was exposed to IR laser radiation for 10 min and antinociceptive effect was determined by means of tail immersion test. The experiments were also performed on 1-NAME and methylene blue pretreated rates, in which both chemicals were administered into right lateral brain ventricle. The results were compared to the ones obtained in the control group in which sham irradiation was made. It was observed that 10 min. exposure to low-power IR laser radiation induced only transient distinct antinociceptive effect in rats. This effect was prevented by ICV. injection of 1-NAME, an inhibitor of nitric oxide synthase and methylene blue, an inhibitor of soluble guanylate cyclase. It seems that nitric oxide is involved in mechanism of low-power laser radiation- induced analgesia.
The aim of the study was to determine the dose-dependent influence of low-power laser radiation on carbohydrate metabolism in 70 male Wistar rats. The animals were primarily divided into 2 groups: B - irradiated group and C - control one in which sham - irradiation was made. The rats from B - group were irradiated daily for 10 minutes with semiconductive laser emitting the radiation of infrared wavelength 904 nm. Within both groups the animals were divided into subgroups (B I - B VII and CI - C VII) in which the dissections were made on 1st, 3rd, 6th, 9th, and 14th day of irradiation and on 5th and 8th day after the end of cycle of irradiation respectively. In all subgroups blood samples were collected to determine the glucose and insulin levels. Parts of the liver and pancreas were taken for histological examination in light microscope and in electron microscope. The lowest, statistically significant glycaemia was observed in the subgroup B V. Significant increase of glycaemia and significantly higher insulin concentration was found only in the subgroup B VI. The I/G ratio increased significantly in the subgroup B V. Lower intensity of paS reaction was presented in subgroups B I, B III, B V, B VI and B VII. The increased amount of paS-positive substances was observed in the I and II zone of liver acinus. Electron microscopic studies of hepatocytes showed: numerous glycogen conglomerations in subgroups B I, B II, B VI and B VII, the extension of RER in B II and B III, light vacuoles in B II, Golgi apparatus and biliary canaliculus expansion in B V and structural changes of several mitochondria - slight swelling or discontinuation of their outer membranes, electron microscopic findings in pancreas cells included: lower number of typical granules in beta and alpha cells as well as Golgi apparatus results it was concluded that the influence of low power laser radiation on carbohydrate metabolism in generally insignificant. It is observed only for higher doses of radiation and does not cause persistent negative effects.
The effect of infrared laser radiation on specific electrical conductivity and relative permittivity of blood in experimental animals was estimated. The experiment was carried out on 30 male guinea pigs weighing 280 - 320 g. Fifteen animals composed a group irradiated with laser, the other 15 animals made up a control group in which no irradiation was made. A semiconductive laser generating infrared radiation (wavelength -- 904 nm, frequency -- 800 Hz, pulse duration -- 200 ns, mean power -- 8.7 mW) was used. The right infracostal region of the animal was irradiated 3 minutes daily for 15 consecutive days. After the end of the experiment in the obtained blood the values of impedance and phase shift angle were measured by means of impedancemeters in the frequency range from 100 to 10,000 kHz. The values of specific electrical conductivity and relative permittivity were calculated on the basis of obtained data. A distinct increase of specific electrical conductivity and decrease of relative permittivity were obtained in the blood of irradiated animals at all frequencies, as compared to the controls. No changes in relaxation frequency were noticed between both groups. On the basis of obtained data it is possible to confirm a distinct effect of low-energy laser radiation on dielectric properties of blood in experimental animals. Since the observed changes in (chi) and (epsilon) ' occurred in (beta) -dispersion region, they might therefore be attributed to the charging of cell membranes in erythrocytes.
The effect on their behavior of chronic exposure to infrared laser radiation applied to the skull of rats was evaluated. The experiment was carried out on 20 Wistar white male rats. A semiconductive infrared laser (wavelength -- 904 nm, frequency -- 100 Hz, pulse duration -- 10 ns, mean power 10 mW, energy density 1.5 J/cm2) was used. A skull fornix of rats was irradiated with use of multidiode probe, 10 minutes daily for 14 consecutive days. A locomotor activity was determined in the `open field' test. Simultaneously, an exploratory activity was examined in the `hole' test. Space memory was determined by means of a water maze test. Afterwards, an irritability was investigated by means of the score of Nakamura and Thoenen. The evaluation of behavior was made 24 hours after a single irradiation, at the 7th and 14th day of repeated everyday irradiation and then at the 7th and 14th day after the end of the cycle of irradiations. Obtained data show that low-energy laser radiation does not affect the rats central nervous system, resulting in persistent changes of behavior.
The influence on pain perception of acute and chronic exposure to infrared laser radiation applied to the skulls of rats is examined. The experiment was carried out on 60 Wistar white male rats. A semiconductive infrared laser (wavelength -- 904 nm, frequency -- 100 Hz, pulse duration -- 10 ns, mean power 10 mW, energy density 1.5 J/cm2) was used. A skull fornix of rats was irradiated with use of a multidiode probe 10 minutes daily for 14 consecutive days. The pain perception was determined by the latency of foot-licking or jumping from the surface of a 56 degree(s)C hot plate. The measurements were made immediately before irradiation, at 1, 5, 10, 15, 20, 25 and 30 minutes after irradiation, and then every 15 minutes until 120 minutes after irradiation. The pain perception is estimated immediately after irradiation, 24 hours after exposure at the 7th and 14th day of irradiation as well as at the 7th and 14th day after the last irradiation. A pretreatment with Naloxone (1 ml/kg of body weight i.p.) was made in a part of the animals in order to evaluate the involvement of the opioid system in the observed effect. Obtained data show that low-energy laser irradiation affects the pain reactivity of rats. The inhibition of the analgesic effect of laser irradiation by the antagonist of opioids -- Naloxone -- and also the prolonged character of this action as a result of direct irradiation of the skull indicate the involvement of the opioid system in the occurrence of laser-induced analgesia.
The effect of infrared laser radiation on viscosity of serum and blood in experimental animals was investigated. The experiment was carried out on 30 male guinea pigs weighing 280 - 320 g. Fifteen animals composed a group irradiated with laser, the other 15 animals made up a control group in which no irradiation was made. A semiconductive laser generating infrared radiation (wavelength -- 904 nm, frequency -- 800 Hz, pulse duration -- 200 ns, mean power - - 8.7 mW) was used. The right infracostal region of the animal was irradiated 3 minutes daily for 15 consecutive days. After the end of the experiment in the obtained blood and serum the measurement of hematocrit and rheologic examination was made in the temperature of 310 K. The estimation of viscosity of whole blood was made by means of low-gradient viscosimeter `low shear,' for shearing velocities: D1 equals 0.116 s-1 and D2 equals 4.59 s-1. The viscosity of serum was estimated using Ubbelohd's microviscosimeter. A significant increase of whole blood viscosity measured for shearing velocity D1 was obtained in irradiated animals as compared to the controls. No statistically significant differences in viscosity of whole blood at clotting speed D2, viscosity of serum and hematocrit were observed between both groups of animals. The obtained data prove a distinct effect of low-energy laser radiation on rheologic properties of blood in experimental animals. The increase of whole blood viscosity, observed only at low shearing velocities, is probably related to the changes of structural and functional properties of cell membranes of erythrocytes resulting in a decrease of aggregation.
It was observed in some experiments that visible laser radiation activates the enzymatic function of mitochondria, while infrared laser radiation affects the enzymatic activity of cellular membranes. The aim of the study was to estimate the activity of some membranous as well as mitochondrial enzymes of hepatocytes in rats irradiated with infrared laser. Experimental material consisted of 38 Wistar rats divided into 2 groups -- a studied group exposed to infrared laser radiation and a control group, in which no irradiation was made. A semiconductive infrared laser (wavelength -- 904 nm, mean power -- 8.9 mW) was used. The clean-shaven skin of the right infracostal region of animals was irradiated 5 minutes daily for 15 consecutive days. After finishing the experiment in the preparations from obtained segments of the left liver lobe, the enzymatic activity of succinate dehydrogenase (SDH, EC 1.3.99.1), lactic dehydrogenase (LDH, EC 1.1.1.27), Mg2+ dependent ATP-ase (ATP-ase Mg2+, EC 3.1.3.2.) and acid phosphatase (AcP, EC 3.6.1.8.) was estimated with the use of histochemical methods. In the case of SDH and LDH the increase of enzymatic activity was observed in all 3 zones of liver cluster, especially in male rats. In the case of ATP-ase Mg2+ and AcP the increase of enzymatic activity in biliary canaliculi of hepatocytes in all zones of the liver cluster was observed. On the basis of the obtained results it was proved that infrared laser radiation activates significantly the enzymatic activity of most of the analyzed enzymes, which means that it affects not only properties of biological membranes but also activates the oxidoreductive processes of organism, as it has been observed for visible laser radiation. On the basis of the spectrum of energetic levels in macromolecules (Jablonski's diagram) the mechanisms of availed results are discussed both for enzymes possessing and not possessing chromatophores.
The neutrophil is the cell in which phagocyting and transforming of some exogeneous agents results in marked stimulation of nonmitochondrial respiratory chain activity (respiratory burst). In our experiment we focused on determining the level of chemiluminescence (CL) of stimulated neurotrophils during and after irradiation, measuring the photon emission intensity in 6 second's intervals. We used Ga-Al-As pulsed laser (wavelength 904 nm, mean power 8,9 mW, Alpha-Electronics GmbH, Germany) which was placed over the tube containing the suspension of guinea pig peritoneal neurotrophils (2X106 cells/ml). The sensitivity range of used photomultiplier (9514s, THORN EMI, Middlesex, England) was 300-600 nm, which allowed us to measure the CL of neutrophils while being irradiated. The neutrophils were stimulated by phorbol myristate acetate (PMA) and CL intensified by luminol. The decay of luminol-dependent CL of neutrophils may be described by hyperbolic function curve. We switched the laser radiation on for 20 s, 60 s and 300 s and each time we observed the same reaction: the about 20% decrease of intensity of CL immediately after beginning the irradiation. The CL remained on decreased level during the whole period of irradiation reaching immediately the level of CL intensity characteristic for decay curve (20% increase), just after switching off the laser. Only after the longest irradiation time (300 s) we observed CL being higher and inconsistent with decay curve for several minutes. The type of reaction was always the same, regardless to the point of CL decay curve at which laser radiation was applied. The same changes of Cl we obtained irradiating the enzymatic system: horseradish peroxidase (HRP)-luminol - H2O2.
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