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Effects of Sucrose and Glycerol during the Freezing Step of Cryopreservation on the Viability of Goat Spermatozoa

  • Farshad, Abbas;Akhondzadeh, S.
    • Asian-Australasian Journal of Animal Sciences
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    • v.21 no.12
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    • pp.1721-1727
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    • 2008
  • Four experiments were conducted to study the following: i) the influence of different concentrations of sucrose (0.15, 0.3 and 0.5 M with osmolality of 308, 500 and 760 mOsm/kg, respectively) in diluents and control diluent (370 mOsm/kg) on intensity of motility and progressive motility of goat sperm without rehydration and freezing step in four incubation periods (0, 0.5, 2 and 4 h after dilution); ii) the influence of gradual dilution (in 3 steps) on improvements in ascertained results of the first experiment; iii) cryoprotective effects of different concentrations of sucrose (0.15, 0.22, 0.29 and 0.37 M with osmolality of 450, 560, 740 and 920 mOsm/kg, respectively) plus 7% glycerol and 20% egg yolk in basic diluent (Tris-Citric acid-Fructose) and iv) the effect of two concentrations of sucrose (0.15 and 0.22 M) with and without glycerol (7%). In experiment 1, we obtained better results for control diluent, 0.15 and 0.3 M sucrose supplemented diluents with 0 and 0.5 h incubation periods. In experiment 2, apart from a slight improvement, similar tendencies to experiment 1 were observed. In experiment 3, we obtained the best result for diluent with 0.22 M sucrose with regard to intensity of motility, progressive motility, live sperm and normal acrosomes ($40{\pm}4%$, $3.1{\pm}0.2$, $37{\pm}4%$ and $37{\pm}4%$, repectively). In experiment 4, we obtained the best result for diluent with 0.22 M sucrose plus 7% glycerol in regard to intensity of motility, progressive motility and live sperm ($39{\pm}3%$, $3.6{\pm}0.4$ and $41{\pm}4%$, respectively). The characteristic normal acrosomes in diluents without glycerol, i.e. diluents with 0.15 and 0.22 M sucrose showed better results ($39{\pm}8$ and $42{\pm}6%$ respectively). With regard to the release of hyaluronidase enzyme there were no significant differences between diluents (p>0.05). The results of the diluents with 0.15 and 0.22 M sucrose without glycerol were slightly lower than those with glycerol ($69{\pm}11$ and $70{\pm}11$ vs. $72{\pm}11$ and $70{\pm}11{\times}120{\times}10^6$ units $ml^{-1}$, respectively). In conclusion, the use of concentrated sucrose solutions showed that goat sperm can tolerate osmolality up to 560 mOsm (0.22 M) in the freezing period. In addition, glycerol proved to be a necessary cryoprotective agent in the cryopreservation of goat sperm, particularly for intensity of motility, progressive motility and live sperm.

Efficiency Calibration of HPGe Detector in Normal ana Coincidence Mode for the Determination of Prompt Gamma-ray (즉발감마선 측정을 위한 HPGe 검출기의 전계수 또는 동시계수모드에서의 광대역 계측효율 보정)

  • 송병철;박용준;지광용
    • Journal of Nuclear Fuel Cycle and Waste Technology(JNFCWT)
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    • v.2 no.2
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    • pp.97-104
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    • 2004
  • Neutron induced prompt gamma-ray spectroscopy(NIPS) system measures the prompt gamma-ray emitting by the interaction of a neutron with various materials. This system will be of great benefit to scientists worldwide, since it provides the non-destructive measurement of many element in either solid or liquid wastes. In this study, the full-energy-peak (FEP) efficiency calibration for a HPGe detector was constructed in the ${\gamma}$-ray energy range from 80 keV to 8 MeV, using $^{l33}$Ba and >TEX>$^{152}Eu$ RI sources and $ ^{35}Cl(n, ${\gamma}$)^{36}Cl$ thermal neutron captured reaction. The FEP efficiency curve for the higher energies using the $^{35}Cl(n, ${\gamma}$)^{36}Cl$ reaction was normalized with the curve obtained from the RI sources, since the accurate activity of its prompt ${\gamma}$-ray is unknown. The average thermal neutron flux was theoretically calculated using the FEP efficiency curve for the KCl standard solutions. The NIPS system equipped with a ${\gamma}$-${\gamma}$ coincidence setup with two n-type coaxial HPGe detectors was considered in order to reduce the interfering ${\gamma}$-ray background. The FEP efficiency curve for the ${\gamma}$-${\gamma}$ coincidence system was also obtained for full energy range. The performance of the normal and coincidence NIPS system was tested by comparing signal-to-noise ratio in each mode using the reference sample.e.

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The Cytoskeletal and Chromosomal Constitution in Vitrified Mouse Oocyte (초자화동결된 생쥐 미수정란의 Cytoskeleton 및 염색체 변화)

  • Yi, B.K.;Kim, E.Y.;Nam, H.K.;Lee, K.S.;Yoon, S.H.;Park, S.P.;Lim, J.H.
    • Clinical and Experimental Reproductive Medicine
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    • v.25 no.3
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    • pp.287-292
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    • 1998
  • The objective of this study was to confirm whether the vitrification method using EFS35 has detrimental effect for cytoskeleton and chromosome constitution of the mouse oocytes by indirect immunocytochemistry and chromosome analysis. Mouse oocytes were vitrified using EFS35 which consisted of 35% ethylene glycol, 18% ficoll, 0.3 M sucrose and 10% FBS in M2 medium. The results obtained in this experiment were summarized as follows: When the survival rates after being exposed or vitrified in EFS35 were examined, there were not different between two groups (97.7 and 89.3%). Also, when the microtubule morphology and microfilament distribution in vitrified oocytes were examined, normal percentage of two cytoskeleton in vitrified group (95.5 and 100%) was not different from that in control (97.5 and 100%) and exposed group (92.3 and 100%). In addition, the rate of oocytes containing a normal chromosome number in vitrified group (73.5%) after IVF was not different from that in control (79.5%) and exposed group (78.7%). These results indicated that the cytoskeletal morphology and chromosome constitution of mouse oocytes were not affected by cryoprotectant (EFS35) or freezing apd that vitrification methods using EFS35 was suitable for cryopreservation of mouse oocytes.

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An Experimental Study on the Changes of Liver Tissue by the Administration of Anticlonorchial Drugs to Rabbits with Clonorchiasis (간(肝)디스토마증(症) 치료약투여후(治療藥投與後) 간장(肝臟) 변화(變化)에 관(關)한 실험적(實驗的) 연구(硏究))

  • Kang, Shin-Wan
    • The Korean Journal of Pharmacology
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    • v.2 no.2
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    • pp.3-22
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    • 1966
  • An experimental study was done on rabbits to observe the effects of several anticlonorchial drugs on the pathology of the liver infested with Clonorchis sinensis. After two months of infestation with Clonorchis sinensis by giving $400{\sim}500$ metacercariae by mouth, hexachlorophene, chloroquine 2,2' methylenebis (3,4,6 trichlorophenoxy acetic acid) and Hetol were administered orally and follow up macro-and microscopic studies of the liver pathology were done in 2 to 3 days, one month, 2 months and 3 months after the completion of medications. The results obtained in this study are as follows: 1. In both groups which were administered hexachlorophene piperazine 20mg/kg for seven days or 8mg/kg for 18 days, the macroscopic findings of the liver after 3 months revealed only minimal changes of the color and consistency The histopathological findings were the reduction of fibrosis, pseudolobulation, proliferation and adenomatous hyperplasia of bile ducts, and regeneration of liver parenchyma. 2. In groups which were administered chloroquine phosphate 20mg/kg for 18 days or 40mg/kg for 5 days, and also in groups which were administered dithiazanine iodide 30mg/kg for 18 days or 60mg/kg for 5 days, no significant findings of recovery were observed either macroscopically or microscopically. 3. In the group which was given 20mg/kg of 2,2' methylenebis(3,4,6 trichlorophenoxy acetic acid) for 5 days, prominent healing of the damaged tissues was observed after 2 months, revealing the decrease of fibrous tissue, caliber of bile ducts and adenomatous hyperplasia of the epitherial cells of the bile ducts, and regenerationof liver parenchyma. 4. In the group which was given Hetol 200mg/kg for 5 days, swelling, congestion and eddish-brown discoloration of the liver were noted macroscopically after 3 days of completion of drug administration. Hemorrhage, congestion, necrosis and degeneration of the parenchyma were observed microscopically After 10 days, liver appeared almost normal macroscopically, but marked fat degeneration was noted microscopically. After 2 months, the liver was almost normal in gross appearence with only slight atrophy and also marked healing was observed microscopically, i. e. decrease of fibrous tissue and reduction of the previously enlarged bile duct and the regeneration of the liver parenchyma.

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Effect of Crude Saponin from Korean Red Ginseng on Clinical Chemical Parameters of Ovariectomized Rat (난소절제 흰쥐의 임상화학지수매 미치는 홍삼조사포닌의 영향)

  • 곽이성;위재준;황석연;경종수;남기열;김시관
    • Journal of Ginseng Research
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    • v.24 no.1
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    • pp.46-50
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    • 2000
  • This study was carried out to investigate the effect of crude saponin from Korean red ginseng on clinical chemical parameters in ovariectomized rats. The crude red ginseng saponin was prepared by Diaion HP-20 adsorption chromatography and spirit. Tota1 of 50 rats were divided into 5 groups: normal control (NC), sham-ovariectomized (SO), ovariectomized (OR), ovariectomized and saponin treated (OS) and normal control treated with saponin (NS). Saponin was intraperitonally administered for 12 weeks since 1 week before ovariectomy: The body weight of ovariectomized rats showed no significant change but that of NS group showed significant increase when compared with NC group. Platelet counts of serum showed significant increase when treated with saponin regardless of ovariectomy. Triglyceride content of serum in NC group was 152.1 mg/㎗, while that of OR group was decreased to 99.9 mg/㎗ However, when saponin was administered, the content was increased to 138.0 mg/㎗. The weight of spleen also showed significant increase when treated with saponin, while the other organs showed no weight changes. On the other hand, ovariectomy in rats induced decrease in femur weight by 10% when compared with NC group. However, administration of crude saponin in ovariectomized rats recovered the weight of the femur to the similar level of NC (e<0.01 0.05). In addition, femur weight of NS group indicated 10 to 16% higher value than that of NC. These results suggest that Korean red ginseng saponin attenuates phyiological disorders induced by malfunction of ovary.

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Development of Moisture Content Prediction Model for Larix kaempferi Sawdust Using Near Infrared Spectroscopy (근적외선 분광분석법을 이용한 낙엽송 목분의 함수율 예측 모델 개발)

  • Chang, Yoon-Seong;Yang, Sang-Yun;Chung, Hyunwoo;Kang, Kyu-Young;Choi, Joon-Weon;Choi, In-Gyu;Yeo, Hwanmyeong
    • Journal of the Korean Wood Science and Technology
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    • v.43 no.3
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    • pp.304-310
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    • 2015
  • The moisture content of sawdust must be measured accurately and controlled appropriately during storage and transportation because biological degradation could be caused by improper moisture. In this study, to measure the moisture contents of Larix kaempferi sawdust, the near-infrared reflectance spectra (Wavelength 1000-2400 nm) of sawdust were used as detection parameter. After acquiring the NIR reflection spectrum of specimens which were humidified at each relative humidity condition ($25^{\circ}C$, RH 30~99%), moisture content prediction model was developed using mathematical preprocessings (e.g. smoothing, standard normal variate) and partial least squares (PLS) analysis with the acquired spectrum data. High reliability of the MC regression model with NIR spectroscopy was verified by cross validation test ($R^2$ = 0.94, RMSEP = 1.544). The results of this study show that NIR spectroscopy could be used as a convenient and accurate method for the nondestructive determination of moisture content of sawdust, which could lead to optimize wood utilization.

A COMPARATIVE STUDY ON THE STRENGTH AND THE BONE FORMATION AT THE RATS CALVARIAL DEFECTS OF DFDB GRAFT AND THOSE OF THE COMPOSITE GRAFT WITH DFDB AND RESORBABLE HYDROXYAPATITE (백서 두개골 결손부에 동결 건조 동종 탈회골을 단일매식한 경우와 동결 건조 동종 탈회골과 흡수성 수산화인회석을 복합매식한 경우의 강도 및 골형성에 관한 비교 연구)

  • Seo, Young-Ho;Yim, Chang-Joon;Lee, Jae-Il
    • Journal of the Korean Association of Oral and Maxillofacial Surgeons
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    • v.26 no.6
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    • pp.557-564
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    • 2000
  • The bone graft materials can be grossly divided into autogenous bone, allogenic bone, xenogenic bone, and alloplastic material. Much care was given to other bone graft materials away from autogenous bone due to its additional operation for harvesting, delayed resorption and limitation of quantity. Demineralized freeze-dried bone(DFDB) and hydroxyapatite are the representatives of bone graft materials. As resorbable hydroxyapatite is developed in these days, the disadvantage of nonresorbability can be overcome. So we planned to study on the strength and the bone formation at the rats calvarial defects of DFDB graft and those of the composite graft with DFDB and resorbable hydroxyapatite. We used the 16 male rats weighting range from 250 to 300 gram bred under the same environment during same period. After we made the 6mm diameter calvarial defect, we filled the DFDB in 8 rats and DFDB and resorbable hydroxyapatite in another 8 rats. We sacrificed them at the postoperative 1 month and 2 months with the periostium observed. As soon as the specimens were delivered, we measured the compressive forces to break the normal calvarial area and the newly formed bone in calvarial defect area using Instron(Model Autograph $S-2000^{(R)}$, Shimadzu, Japan). The rest of the specimens were stained with H&E(Hematoxylin & Eosin) and evaluated with the light microscope. So we got the following results. 1. In every rats, there was no significant difference between the measured forces of normal bone area and those of the bone graft area. 2. In 1 month, the measured forces at DFDB graft group were higher than those of the DFDB and resorbable hydroxyapatite composite graft group(P<0.05). 3. In 2 months, there was no significant differences between the measured forces of DFDB graft group and those of the DFDB and resorbable hydroxyapatite composite graft group. 4. In lightmicroscopic examination, most of the grafted DFDB were transformed into bone in 1 month and a large numbers of hydroxyapatite crystal were observed in DFDB and resorbable hydroxyapatite composite graft group in 1 month. 5. Both group showed no inflammatory reaction in 1 month. And hydroxyapatite crystals had a tight junction without soft tissue invagination when consolidated with newly formed bone. 6. In both groups, newly formed bone showed the partial bone remodeling and the lamellar bone structures and some of reversal lines were observed in 2 months. From the above results, it is suggested that DFDB and resorbable hydroxyapatite composite graft group had a better resistance to compressive force in early stage than DFDB graft group, but there would be no significant difference between two groups after some period. And it is suggested that the early stage of bone formation procedure of DFDB and resorbable hydroxyapatite composite graft group was slight slower than that of DFDB graft group, but there would be no significant difference between two groups after some period.

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Effects of Mahwangbujaseshin-tang (Mahuangfuzixixintang) (麻黃附子細辛湯) on MIA-Induced Osteoarthritis Rats (마황부자세신탕(麻黃附子細辛湯)이 MIA로 유도된 골관절염 유발 Rat에 미치는 영향)

  • Lee, Hyung-Eun;Oh, Min-Seok
    • Journal of Korean Medicine Rehabilitation
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    • v.24 no.2
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    • pp.65-81
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    • 2014
  • Objectives This study was carried out to find out the anti-osteoarthritic effects of Mahwangbujaseshin- tang (Mahuangfuzixixintang ) on the monosodium iodoacetate (MIA)-induced osteoarthritis rats. Methods Osteoarthritis was induced by injecting MIA ($50{\mu}l$) into the knee joint of rats. Rats were divided into a 3 groups (n=7). The injection did not fit the normal group. A week later, after the injection of MIA, while control group took normal saline 2 ml, the extract of Mahwangbujaseshin-tang (Mahuangfuzixixintang ) (MBST) (200 mg/kg) was injected to treated group. After that, we examined hind paw weight bearing ability, functions of liver and kidney, serum TNF-$\alpha$, IL-$1{\beta}$, IL-6, $PGE_2$, $LTB_4$, TIMP-1, MMP-9 and hematology. Volume of cartilage was measured by micro CT arthrography. Injury of synovial tissue was measured by H & E, Safranin-O immunofluorescence. Results 1) DPPH and ABTS free radical scavenging activity of MBST was increased according to concentration of MBST and total phenolic contents were in high level. 2) In RAW 264.7 cells, ROS production was significantly decreased in MBST (at 10, $100{\mu}g/ml$) and NO was also decreased but meaningless in MBST (at $100{\mu}g/ml$). 3) In RAW 264.7 cells, IL-6 production was significantly decreased in MBST (at $100{\mu}g/ml$) and TNF-$\alpha$ and IL-$1{\beta}$ production were also decreased but meaningless in MBST (at $100{\mu}g/ml$). 4) In hind legs weight-bearing measurement, level of weight-bearing was increased. 5) Functions of liver and kidney were not affected. 6) TNF-$\alpha$, IL-$1{\beta}$, IL-6, $PGE_2$, $LTB_4$, MMP-9 and TIMP-1 production were significantly decreased. 7) In hematology, the levels of neutrophils, monocytes were significantly decreased and the levels of white blood cells, lymphocytes were also decreased but meaningless. 8) In micro CT-arthrography, cartilage volume was significantly increased. 9) Histopathologically, injury on cartilage and synovial membrane of MBST group was decreased. Conclusions Based on all results mentioned above, Mahwangbujaseshin-tang (Mahuangfuzixixintang) is believed to be meaningful for suppressing the progress of osteoarthritis. And it is related to inhibiting the activity of inflammatory cytokine and injury of volume in cartilage.

A STUDY OF THE DEVELOPMENT AND STANDARDIZATION OF ADHD DIAGNOSTIC SYSTEM (전산화된 주의력장애 진단시스템의 개발 및 표준화 연구)

  • Cho, Sung-Zoon;Chun, Sun-Young;Hong, Kang-E;Shin, Min-Sup
    • Journal of the Korean Academy of Child and Adolescent Psychiatry
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    • v.11 no.1
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    • pp.91-99
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    • 2000
  • Objectives:Present study developed the computerized ADHD Diagnostic System(ADS) in order to diagnose ADHD and evaluate treatment effect of it, and conducted a standardization study for ADS. Methods:The normative group was composed of 847 children and adolescents between the age of 5 and 15(boy 429, girl 418) living in the areas of Seoul, Kyunggi-do, and Kangwon-do. 30 ADHD children with age ranged 7 to 9 years were participated present study to evaluate the validity of ADS. To establish the norms for diagnosing ADHD, the means and standard deviations of normative group were used to calculate T-scores for each age group. Results:The reliability coefficient of ADS(Cronbach's ${\alpha}$) was .85. There were significant differences in the measures of ADS except commission error between the normal and the ADHD groups. Three factors were extracted through factor analysis of ADS, which were labelled 'inattention', 'slow information processing' and 'impulsivity'. Discriminant analysis showed that ADS significantly discriminate the normal and the ADHD groups. Percentage of correct classification by ADS variables was 96.7%. Conclusion:These results put together strongly support the reliability and validity of ADS as a diagnostic instrument for ADHD.

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Analysis of Immunoglobulin λ Light Chain Repertoire in Systemic Lupus Erythematosus (루푸스 환자의 면역글로불린 λ 경쇄 레파토리 분석)

  • Chang, Ji Eun;Lee, Jisoo
    • IMMUNE NETWORK
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    • v.3 no.3
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    • pp.227-234
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    • 2003
  • Background: Immunoglobulin (Ig) light chain repertoire has been implicated as a critical determinant in regulation of autoreactive B cells and production of pathogenic anti-DNA antibodies in systemic lupus erythematosus (SLE). We analyzed the impact of Ig ${\lambda}$ chain repertoire on development of autoimmunity in patients with SLE. Methods: We obtained genomic DNA from individual peripheral CD19+ B cells of 3 untreated active SLE patients, and amplified $V{\lambda}$ rearrangements from each single cell by polymerase chain reaction. Results: A total number of 208 $V{\lambda}J{\lambda}$ rearrangements were analyzed. Analyzed sequences included 158 productive rearrangements and 50 nonproductive rearrangements. The differences in $V{\lambda}$ gene usage in the productive and nonproductive repertoire of SLE patients were found compared to the non-autoimmune individuals. $V{\lambda}$ gene, 9A was significantly overrepresented in nonproducative repertoire of SLE patients (P=0.016). In the productive repertoire, $V{\lambda}$ genes, 3L and 1E were found more often in the SLE patients (P=0.001, P=0.043). When the productive and the nonproductive repertoires were compared, 9A was found significantly less in the productive repertoire in the SLE patients (P=0.000). There were no significant differences in the $J{\lambda}$ gene usage between SLE patients and non-autoimmune individuals, but $J{\lambda}2/3$ gene was the most frequently used in SLE, whereas $J{\lambda}7$ gene was the most frequently used in the normal subjects. In the productive SLE $V{\lambda}$ repertoire, 9.4% of the total sequences employed identical CDR3. It was particularly striking to find 7 identical versions of the 1G-$J{\lambda}2/3$ $V{\lambda}J{\lambda}$ rearrangements from one patient and 3 of the same sequence from another patient. Notably, identical $V{\lambda}$ junctions in the SLE patients utilized significantly more homologous joining compared to $V{\lambda}$ junctions of the normal adults (P=0.044). Conclusion: These data demonstrate regulation of ${\lambda}$ light chain expression in the SLE patients by selection of unique $V{\lambda}$ genes. Also, biased selection and clonal expansion of particular $V{\lambda}$ rearrangements are apparent in the SLE ${\lambda}$ repertoire.