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Effect of Cryoprotectant Kinds and Cell Stages on the Viability of Mouse Embryos Cryopreserved by OPP Vitrification (동결보호제의 종류 및 배발달단계가 OPP Vitrification 동결보존시 생쥐수정란의 생존성에 미치는 영향)

  • 공일근;조성균;조성근
    • Korean Journal of Animal Reproduction
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    • v.23 no.1
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    • pp.85-92
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    • 1999
  • This study was designed to determine effect of cryoprotectant kinds and cell stages on OPP vitrification method in mouse embryos. The freezing speed, cryoprotectants and cell stage could affect of embryo viability following various vitrification methods. The vitrification solution used were consisting of 40% (v/v) ethylene glycol, 18% (w/v) Ficoll, 0.3 M sucrose solution in holding medium (D-PBS supplemented with 5% FCS: HM) (EFS) or 16.5% ethylene glycol , 16.5% dimethyl sulfoxide, 0.5 M sucrose in HM (EDS). The embryos were collected from oviduct at 18 h after hCG injection and then washed and cultured in mHTF medium until use. In experiment 1, the blastocysts were vitrified by OPP straw to determine the optimal vitrification solution of EFS or EDS. The post-thaw survival rates at re-expanded stage rates were significantly different between EFS and EDS (95.0 vs 100%), but at hatching stage was not different between EFS and EDS (90.0 vs 95.0%). respectively. In experiment 2, zygotes, 2-, 4-cell, morula and blastocysts were vitrified by OPP method to determine the acceptable of early stage embryos. The development rates to expanded blastocyst in zygote (70.0%) were significantly lower rather than those in 2-, 4- 8-cell, compacted morula or blastocyst (89.7, 90.0, 92.8, 97.6 or 97.5%), respectively. However, the cell number of post-thaw developed to expanded blastocyst in blastocyst and control blastocyst stage (39.6$\pm$2.81, 35.7$\pm$2.98) were significanty higher than those in zygote, 2-, 4-, 8-cell, compacted morula (29.8$\pm$3.21, 31.3$\pm$3.83, 29.3$\pm$3.58, 28.9$\pm$3.21 or 30.8$\pm$2.93). In experiment 3, the zygotes were exposed in VSl for 1, 2, and 3 min to the optimal exposed time. The cleavage rates (91.6, 88.5, 88.9%) and develop mental rates to blastocyst (83.3, 74.3 and 69.4%) depends on the exposed time in VSl were not significantly different among 1, 2, or 3 min, respectively. The cell number also were not significantly different among exposed time in VS1. respectively. These results indicate that OPP method could be useful for vitrification either EFS or EDS vitrification solution. The post-thaw survival rates at zygote were significantly lower than those at 2-, 4-, 8-cell, morula or blastocyst, respectively. The zygote stage were more sensitive rather than late stage embryos. The exposing time in VS1 for 1 min was better than that for 2 or 3 min, even it was not significantly different. The OPP vitrification method could be useful of mouse embryos either with EFS or EDS vitrification solution.

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Comparisons of occlusal force according to occlusal relationship, skeletal pattern, age and gender in Koreans (한국인에서의 부정교합 여부와 골격형태, 연령, 성별에 따른 교합력의 비교)

  • Yoon, Hye-Rim;Choi, Yoon-Jeong;Kim, Kyung-Ho;Chung, Choo-Ryung
    • The korean journal of orthodontics
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    • v.40 no.5
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    • pp.304-313
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    • 2010
  • Objective: The aim of this study was to evaluate the occlusal force and contact area and to find its associating factors in Koreans. Methods: Occlusal force and contact area in maximum intercuspation were measured using the Dental $Prescale^{(R)}$ system in 651 subjects (15 with normal occlusion, 636 with various malocclusions divided into subgroups according to the skeletal pattern, Angle's molar relationship, age and gender). Results: Occlusal force of the normal occlusion group ($744.5{\pm}262.6N$) was significantly higher than those of the malocclusion group ($439.0{\pm}229.9N$, $p$ < 0.05). Occlusal force was similar regardless of differences in ANB angle or Angle's molar classification, however the increase in vertical dimension significantly reduced occlusal force ($p$ < 0.05). Conclusions: Occlusal force was significantly lower in the malocclusion group compared to the normal occlusion group, and in females compared to males, but it was not affected by age, antero-posterior skeletal pattern or molar classification. Although a hyperdivergent facial pattern indicated lower occlusal force compared to a hypodivergent facial pattern, the differences in skeletal pattern were not the primary cause of its decrease, but a secondary result induced by the differences in occlusal contact area according to the facial pattern.

Effects of Supplementary Multiple Probiotics or Single Probiotics on the Performance, Intestinal Microflora, Immune Response of Laying Hens and Broilers (혼합 또는 단일 생균제가 산란계와 육계의 생산성, 소장내 미생물 균총 및 면역 체계에 미치는 영향)

  • Kim, Chan-Ho;Woo, Kyung-Chun;Kim, Geun-Bae;Park, Yong-Ha;Paik, In-Kee
    • Korean Journal of Poultry Science
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    • v.37 no.1
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    • pp.51-62
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    • 2010
  • This study was conducted to investigate the effects of dietary supplementation of multiple probiotics on the performance, small intestinal microflora and immune response in laying hens and broilers. In Exp.1, a total of 800, 82 wk old Hy-line Brown$^{(R)}$ laying hens were assigned to one of the following five dietary treatment; Control, Antibiotics (avilamycin 6 ppm), Probiotics; PB-M (Micro-ferm$^{(R)}$ 0.2%), PB-L (Lacto-sacc$^{(R)}$ 0.1%), PB-Y (Y University probiotics 0.2%). Each treatment was replicated eight times with 20 birds in each replicate and two birds were housed in each cage. Twenty birds units were arranged according to completely randomized block design. Feeding trial lasted 6 wk under 16 h lighting regimen. The Exp. 2, was conducted with a total of 1,000 broilers chicks (Ross$^{(R)}$). They were divided into five treatments, same as those of Exp. 1. Birds were fed starter (0~3 wk) and grower (4~5 wk) diets. Each treatment was replicated four times with 50 birds per pen comprising of deep litter. In Exp. 1, egg production parameters, such as hen-day and hen-house egg production, egg weight, broken and soft shell egg production, feed intake and feed conversion were not significantly different among treatments. However, strength and thickness of eggshell were significantly (P<0.05) different. Among the probiotics, PB-Y showed the highest strength and thickness of eggshell. Eggshell color, egg yolk color and Haugh unit were not significantly influenced. In Exp. 2, overall weight gain (0~5 wk) and mortality were not significantly different among treatments. However, weight gain of birds from PB-Y treatment during starter (0~3 wk) was significantly lower than the birds from Control and Antibiotic treatment. During the whole period (0~5 wk), birds from Antibiotics treatment had higher feed intake and Production Index (PI) and lower feed conversion than birds from Control treatment. Probiotics treatments were not significantly different from the Control on feed intake and feed conversion. In Exp.1, there were significant (P<0.05) differences in leukocytes parameters, such as white blood cell (WBC), hetrophil (HE), lymphocytes (LY), monocyte (MO), eosinophil (EO) and stress index (SI; HE/LY) in the blood of layers. Birds from Antibiotics and probiotics treatments tended to increase these parameters. In Exp. 2, however, only SI was significantly (P<0.05) decreased in Antibiotics treatments. Concentration of serum immunoglobulin (IgG) were higher (P<0.05) in PB-M and PB-Y treatments when compared with Control treatment in Exp. 1. The population of E. coli significantly (P<0.05) decreased in birds from Antibiotics, PB-L and PB-Y treatments when compared with birds from Control treatment in Exp. 1. Metalbolizability of crude fat decreased significantly (P<0.05) in birds from probiotic treatments in Exp. 2. It was concluded that the response of probiotics on the productivity of layers and broilers were different. Probiotics increased strength and thickness of eggshell in layers, and decreased feed conversion and increased PI in broilers. Leukocytes and IgG tended to increase by supplementation of antibiotics and probiotics in layers. Intestinal E. coli tended to decrease in layers. Digestibility of crude fat of diet decreased in probiotics treatments broilers. Parameters of blood and microbial were more sensitive in layers than broilers.

The Risk Assessment of Butachlor for the Freshwater Aquatic Organisms (Butachlor의 수서생물에 대한 위해성 평가)

  • Park, Yeon-Ki;Bae, Chul-Han;Kim, Byung-Seok;Lee, Jea-Bong;You, Are-Sun;Hong, Soon-Sung;Park, Kyung-Hoon;Shin, Jin-Sup;Hong, Moo-Ki;Lee, Kyu-Seung;Lee, Jung-Ho
    • The Korean Journal of Pesticide Science
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    • v.13 no.1
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    • pp.1-12
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    • 2009
  • To assess the effect of butachlor on freshwater aquatic organisms, acute toxicity studies for algae, invertebrate and fishes were conducted. The algae grow inhibition studies were carried out to determine the growth inhibition effects of butachlor (Tech. 93.4%) in Pseudokirchneriella subcapitata (formerly knows as Selenastrum capriconutum), Desmodesmus subspicatus (formerly known as Scendusmus subspicatus), and Chlorella vulgaris during the exposure period of 72 hours. The toxicological responses of P. subcapitata, D. subspicatus, and C. vulgaris to butachlor, expressed in individual $ErC_{50}$ values were 0.002, 0.019, and $10.4mgL^{-1}$, respectively and NOEC values were 0.0008, 0.0016, and $5.34mg\;L^{-1}$, respectively. P. subcapitata was more sensitive than any other algae species. Butachlor has very high toxicity to the algae, such as P. subcapitata and D. subspicatu. In the acute immobilisation test for Daphnia magna, the 24 and $48h-EC_{50}$ values were 2.55 and $1.50mg\;L^{-1}$, respectively. As the results of the acute toxicity test on Cyprinus carpio, Oryzias latipes and Misgurnus anguillicaudatus, the $96h-LC_{50}s$ were 0.62, 0.41 and $0.24mg\;L^{-1}$, respectively. The following ecological risk assessment of butachlor was performed on the basis of the toxicological data of algae, invertebrate and fish and exposure concentrations in rice paddy, drain and river. When a butachlor formulation is applied in rice paddy field according to label recommendation, the measured concentration of butachlor in paddy water was $0.41mg\;L^{-1}$ and the predicted environmental concentration (PEC) of butachlor in drain water was $0.03 mg\;L^{-1}$. Residues of butachlor detected in major rivers between 1997 and 1998 were ranged from $0.0004mg\;L^{-1}$ to $0.0029mg\;L^{-1}$. Toxicity exposure ratios (TERs) of algae in rice paddy, drain and river were 0.004, 0.05 and 0.36, respectively and indicated that butachlor has a risk to algae in rice paddy, drain and river. On the other hand, TERs of invertebrate in rice paddy, drain and river were 3.6, 50 and 357, respectively, well above 2, indicating no risk to invertebrate. TERs of fish in rice paddy, drain and river were 0.58, 8 and 57, respectively. The TERs for fish indicated that butachlor poses a risk to fish in rice paddy but has no risk to fish in agricultural drain and river. In conclusion, butachlor has a minimal risk to algae in agricultural drain and river exposed from rice drainage but has no risk to invertebrate and fish.

Changes in Serum IGF-I and Spermatogenesis Analysed by Flow Cytometry in Growing Male Rabbit (성장 중인 수토끼에서 혈청 IGF-I 수준과 Flow Cytometry 측정에 의한 정자 형성의 변화)

  • Lee J. H.;Kim C. K.;Chang Y. M.;Ryu J. W.;Park M. Y.;Chung Y. C.;Pang M. G.
    • Reproductive and Developmental Biology
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    • v.29 no.3
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    • pp.163-168
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    • 2005
  • The aim of this study was to investigate the changes in insulin-like growth factor-I (IGF-I) and growth hormone (GH) in serum, the quantitation of spermato-genesis and the comparable relationships among these measurements during pubertal period in New Zealand White male rabbits. To investigate the age-related testicular changes in DNA contents of spermatogenic cells, the fine-needle testicular biopsies from males aged 10 to 28 wks were evaluated by flow cytometry(FCM). Body weight increased significantly between the ages of 12 and 20 wks (P<0.05) and reached 3.4 kg at 28 wks of age. The highest serum IGF-I level (451.3ng/mL) was observed at 20wks of age (P<0.05) and thereafter remained stable at low levels. Serum GH level at 18 wks of age was 183.3 pg/mL which was significantly higher compared to the other ages (P<0.05), and the rising time in serum GH tend to be somewhat earlier than that of IGF-I. The relative percentage of It-cells in testicular cell compartments was $48.2\%$ at the age of 18 wks which significantly increased than those of 16-wk-old (P<0.05) and thereafter increased with the advance of age to $68\%$. The percentage of 2C-cells in testis was $26.8\%$ at 18 wks of age which was significantly lower than $54.3\%$ at 16 wks old (P<0.05). The percentage of 4C-cells was constantly maintained $2\~6\%$ except the $9.9\%$ at 18 wks of age. In conclusion, the results suggest that the puberty onset occurred at about the 18 wks of age and that the IGF-I and GH in serum during the pubertal period showed the age/growth-specific changes and these changes might be related to the spermatogenesis. The DNA FCM combined with fine-needle testicular biopsy could offer a very sensitive method to monitor the quantitative spermatogenic events related to the puberty onset.

The Nasal Airflow Pressure Monitoring and the Measurement of Airway Pressure Changes in Obstructive Sleep Apnea Syndrome and Upper Airway Resistance Syndrome (수면무호흡증과 상기도저항 증후군에서 Nasal Airflow의 압력측정 및 상기도 압력변화에 대한 연구)

  • Kim, Hoo-Won;Hong, Seung-Bong
    • Sleep Medicine and Psychophysiology
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    • v.7 no.1
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    • pp.27-33
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    • 2000
  • Objectives: The sensitivity and accuracy of thermistor airflow signal has been debated. The purposes of this study were to compare apnea-hypopnea index(AHI) detected from a conventional thermistor signal and a nasal pressure transducer of airflow(NPT), to evaluate the value of NPT for the diagnosis of upper airway resistance syndrome(UARS), and to measure airway pressure fluctuations which produced respiratory arousals in UARS by naso-oro-esophageal manometer catheter. The subjects were 30 patients with obstructive sleep apnea syndrome [mild(540), 10), and 6 UARS patients. Airway resistance arousal in this study was defined as arousals which were not associated with apnea or hypopnea of thermistor signal, but showed significant decrease of nasal airflow pressure just before arousal and a prompt recovery of nasal airflow pressure after arousal. The airway pressure fluctuations were measured during 260 airway resistance arousals observed in 10 patients with OSAS, 2 with UARS. Results: Mean AHIs of patients with OSAS were 33.4 by thermistor and 48.4 by NPT. The AHIs of mild, moderate and severe OSAS groups were 10.2, 32.1, 65.4 respectively by thermistor and 23.1, 45.9, 76.4 by NPT. The mean AHI of patients with UARS was 3.2 by thermistor and 10.8 by NPT. The mean AHI of patients with nonspecific arousals was 2.7 by thermistor and 4.4 by NPT. The mean airway pressure changes during respiratory arousals of different groups were $8.7\;cmH_2O$ in mild OSAS, $11.4\;cmH_2O$ in moderate OSAS, $24.7\;cmH_2O$ in severe OSAS and $6.6\;cmH_2O$ in UARS. Conclusion: The nasal pressure transducer of airflow was more sensitive and accurate for assessing respiratory disturbances of patients with OSAS and was extremely helpful for the diagnosis of UARS without esophageal pressure monitoring. From the results, we would like to propose carefully the NPT diagnostic criteria for sleep disordered breathing as follows: NPT-AHI 5-15 $\rightarrow$ UARS, 15-35 $\rightarrow$ mild OSAS, 35-55 $\rightarrow$ moderate OSAS and >55 $\rightarrow$ severe OSAS.

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THE EFFECTS OF DRYING AGENTS AND BONDING AGENTS ON THE SHEAR BOND STRENGTH OF SEALANTS TO ENAMEL (치면건조제와 접착제의 사용에 따른 치면열구전색재의 전단결합강도에 관한 연구)

  • Lim, Hyun-Hwa;Jang, Ki-Taek;Kim, Chong-Chul;Hahn, Se-Hyun
    • Journal of the korean academy of Pediatric Dentistry
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    • v.30 no.2
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    • pp.196-203
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    • 2003
  • The application of sealants is a highly technique-sensitive procedure, requiring an extremely dry field prior to placement. Moisture contamination of the etched enamel surface before sealant placement is cited as the main reason for sealant failure. The purpose of this study was to evaluate the effects of different methods of sealant application on the shear bond strength of sealants to enamel. In groups 1, 2, 3, 4 Teethmate(unfilled sealant) was used, while Ultraseal XTplus(filled sealant) was used in groups 5, 6, 7, 8. Groups 1 and 5(control) were acid etched for 15 seconds using 35% phosphoric acid, washed and then dried. In groups 2, 6 drying agents were applied, and in groups 3, 7 bonding agents were applied and light cured. In groups 4 and 8 both drying agent and bonding agent were applied. Then sealant was cured to the specimen using molds 3mm in diameter and 2mm in height. Thermocycling was performed and shear bond strength was finally measured. The following results were obtained : 1. Groups using filled sealant(groups 5, 6, 7, 8) showed higher shear bond strengths compared to groups using unfilled sealant(groups 1, 2, 3, 4). 2. Among groups using unfilled sealant(groups 1, 2, 3, 4), groups 2, 3, 4 showed significantly higher shear bond strength compared to group 1(p<0.05). There were no significant differences among groups 2, 3 and 4. 3. There were no significant differences(p>0.05) among groups using filled sealant(groups 5, 6, 7, 8). 4. When modes of fracture were examined, cohesive failure was observed in groups 2, 3 and 4.

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Liquid Chromatography Quadrupole Time-Of-Flight Tandem Mass Spectrometry for Selective Determination of Usnic Acid and Application in Pharmacokinetic Study

  • Fang, Minfeng;Wang, Hui;Wu, Yang;Wang, Qilin;Zhao, Xinfeng;Zheng, Xiaohui;Wang, Shixiang;Zhao, Guifang
    • Bulletin of the Korean Chemical Society
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    • v.34 no.6
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    • pp.1684-1688
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    • 2013
  • A rapid and sensitive method for determining usnic acid of Lethariella cladonioides in rat was established using high performance liquid chromatography (HPLC) quadrupole time-of-flight (QTOF) tandem mass (MS/MS). Rat plasma was pretreated by mixture of acetonitrile and chloroform to precipitate plasma proteins. Chromatographic separation was achieved on a column ($50{\times}2.1$ mm, $5{\mu}m$) with a mobile phase consisting of water (containing $5{\times}10^{-3}$ M ammonium formate, pH was adjusted to 3.0 with formic acid) and acetonitrile (20:80, v/v) at a flow rate of 0.3 mL/min. A tandem mass spectrometric detection with an electrospray ionization (ESI) interface was conducted via collision induced dissociation (CID) under negative ionization mode. The MS/MS transitions monitored were m/z 343.0448 ${\rightarrow}$ m/z 313.2017 for usnic acid and m/z 153.1024 ${\rightarrow}$ m/z 136.2136 for protocatechuic acid (internal standard). The linear range was calculated to be 2.0-160.0 ng/mL with a detection limit of 3.0 pg/mL. The inter- and intra-day accuracy and precision were within ${\pm}7.0%$. Pharmacokinetic study showed that the apartment of usnic acid in vivo confirmed to be a two compartment open model. The method was fully valid and will probably be an alternative for pharmacokinetic study of usnic acid.

Simultaneous determination of illegal galactagogue adulterants in supplement diets by LC-MS/MS

  • Lee, Ji Hyun;Cho, So-Hyun;Park, Han Na;Park, Hyoung Joon;Kim, Nam Sook;Park, Sung Kwan;Kang, Hoil
    • Analytical Science and Technology
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    • v.31 no.4
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    • pp.171-178
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    • 2018
  • Recently, for successful lactation, many breastfeeding mothers seek various products, including herbal medicine, dietary supplements, and prescribed medicines, to improve milk production. As demand for galactogogues grows, it is highly possible that pharmaceutical galactogogues may be adulterated with illegal products to maximize their efficacy. For continuous control and supervision of illegal products, we developed and validated a simple and sensitive LC-MS/MS method capable of simultaneously determining five galactogogues. Chromatographic separation was conducted using an Agilent Poroshell $120SB-C_{18}$ column with a mobile phase consisting of 20 mM ammonium formate (pH 5.4) and 100 % acetonitrile. The total run time was 13 min per analyte. The proposed method was performed according to the guidelines of the International Conference of Harmonization and it produced reliable results. This method showed high sensitivity and specificity, with a limit of detection (LOD) and limit of quantitation (LOQ) of 0.01-0.82 ng/mL and 0.02-2.45 ng/mL, respectively, for the solid- and liquid-type samples. Specificity was evaluated by analyzing matrix-blank samples spiked with the target compounds at LOQ levels, which provided a good separation of all peaks without interference. Additionally, the repeatability and intermediate precision were typically <15 %, whereas the recovery was 80-120 % of the values obtained using blank samples. Thus, we concluded that this method could be used for the identification and quantification of galactogogues in food or herbal products.

The Second-order Scattering of the Interaction of Pd Nanoparticles with Protein and Its Analytical Application

  • Guo, Xiaoyan;He, Baolin;Sun, Chuntao;Zhao, Yanxi;Huang, Tao;Liew, Kongyong;Liu, Hanfan
    • Bulletin of the Korean Chemical Society
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    • v.28 no.10
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    • pp.1746-1750
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    • 2007
  • The second-order scattering (SOS) phenomenon of the interaction of Pd nanoparticles with protein was reported and a simple, sensitive, palladium nanoparticle-based assay for trace amount of protein with SOS technique was developed. The SOS intensities were significantly enhanced due to the interaction of Pd nanoparticles with bovine serum albumin (BSA) or human serum albumin (HSA) at pH 3.5 or 4.0, respectively. The maximum SOS peak appeared at 260/520 nm (λex/λem). The optimal experiment conditions, affecting factors and the influence of some coexisting substances were checked. The SOS intensity increased proportionally with the increase of Pd concentration below 3.0 × 10?5 mol·L?1, while declined gradually above 4.0 × 10?5 mol·L?1. BSA within the range of 0.01-2.6 μg·mL?1 and HSA of 0.01-1.7 μg·mL?1 can be detected with this method and the detection limits were 2.3 and 11.2 ng·mL?1, respectively. The method was successfully applied to the quantitative detection of total protein content in human serum samples with the maximum relative standard deviation (RSD) lower than 2.6% and the recoveries over the range of 99.5-100.5%.