Indonesian Journal of Biotechnology
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Cortisol and Estradiol Profile in Cross-bred Ettawa Does: The Effects of Body Condition Scoring (BCS).
No full textBody Condition Scoring (BCS) is an estimation of the muscle and fat development of an animal. Thin ewes that are fighting to maintain their own body weight and low concentration of cortisol are not able to ovulate as ewes in a more desirable condition due to lack of oestradiol concentration. The aims of this research are to monitor the cortisol and oestradiol profile in Cross-bred ettawa does and to determine effect of BCS on the cortisol and oestradiol profile. Eight does were used in this research. These animals were devided equally into 2 groups based on Body Condition Scoring (BCS), namely BCS 2, which body weight range between 25-30 kgs as group I ( >n=4 ) and BCS 3 which consists of ettawa with body weight range between 33-40 kg as group II ( n=4 ). All animals were synchronized using implant of CIDR and PGF2alpha. Blood from jugular vein were collected every 3 and 6 hours as soon as oestrus until 72 hours. Serum contained cortisol and oestradiol then assayed using ELISAmethod. Cortisol and oestradiol concentrations were compared between groups by T test. The results showed that average concentration of cortisol is 47.17 ± 42.19 ng/mL for BCS 2 and 112.40±74.41ng/mL for BCS 3 (P<0.05), whereas concentration of oestradiol is 72.25±30.62 pg/mL for BCS 2 and 145.72±100.18 pg/mL for BCS 3 (P<0.05). Either cortisol or oestradiol have very synchronized wave except 2 of animals from BCS 2 (50%), which has tendency to suppress each other. It was concluded that profile of cortisol and oestradiol hormone have a very similar pattern, and BCS can affect hormone profile
Epitope Mapping of Fc gamma RIIa Monoclonal Antibodies
No full textFcγRIIa (CD32) is an IgG receptor which has been shown to be important in autoimmune disease pathology. IV.3, 8.7, and 7.30 are anti-FcγRIIa monoclonal antibodies (mAbs), which block the interaction between FcγRIIa and complex IgG. In this study, the three mAbs were demonstrated to inhibit FcγRIIa function. The determination of the precise epitopes of the IV.3, 8.7, and 7.30 mAbs may become a potential approach for designing inhibitors for FcγRIIa. The epitope of IV.3, 8.7, and 7.30 were determined using chimeric receptors based on the extracellular domains of FcγRIIa and the FcεRI a chain. The epitopes for IV.3 was found to be mapped on amino acid residues 132-137, while 8.7 and 7.30 were on amino acid residues 112-119 and 157-162. Based on the crystal 3D model of FcγRIIa molecule, these amino acid sequences are clustered together forming a contiguous region within the ligand binding site of the receptor
Nuclear Maturation of Porcine Oocytes in vitro: Effect of the Cumulus-Oocyte Complexes Quality
No full textThe objective of this study was to examine the effect of the cumulus-oocyte complexes (COCs) quality on the ability of porcine oocytes to mature in vitro. Porcine COCs were collected from 2-6 mm follicles of slaughterhouse ovaries. The oocytes used for IVM were classified into three categories based on the compactness and transparency of the cumulus investment and homogeneity and transparency of the ooplasm. The oocytes were then matured in vitro for 44 h. At 22 of maturation culture, most of the oocytes in allgroups were identified still at germinal vesicle (GV) stage and metaphase I (M-I) stage. After 44 h of culture, a greater proportion of Category I and II oocytes completed in vitro maturation through the second meiotic as compared with that of Category III oocytes (P<0.05). The proportion of oocytes remaining at M-I stage and the degenerative oocytes in Category III oocytes were significantly higher than those of oocytes in other groups (P<0.05). These data indicate that porcine oocytes with high quality cytoplasm and a cumulus cell complement have a much greater chance of maturing in vitro than that lower quality oocytes. The morphological grading of immature oocytes is an appropriate selection criterion for their developmental ability
Actin Distribution in Lamina Neuralis During Cranial Neurulation of Wistar Rats Embryo (Rattus rattus)
No full textsuch as craniorachisis and exencephaly. One of the processes is changing in lamina neuralis cells shape, which iscaused by actin microfilament rearrangement within lamina neuralis cells. To examine the distribution of actinmicrofilament during cranial neurulation Wistar rats embryo were used. Embryos were obtain at following days ofdevelopment; 8 days 18 hours, 9 days, 9 days 6 hours, 9 days 12 hours, 9 days 18 hours, and 9 days 20 hoursrespectively. Immunohistochemistry Avidin Biotin-peroxidase Complex (ABC) method was used to examine andidentify the distribution of actin in lamina neuralis cells. Light microscopic observation shows positive reaction foractin immunoreactivity in the apical surface of bending lamina neuralis cells. In contrast, actin is not observed in nonbendinglamina neuralis. Actin is not detected at 8 days 18 hours embryos. At 9 days embryos, positive reaction isobserved over the entire apical surface of lamina neuralis.Key words: Cranial neurulation, Actin, lamina neuralis, Rats embryo
Identification of Pathogenecity of Avian Influenza Virus Subtype H5N1 from Waterfowls Base on Amino Acid Sequence of Cleavage Site Hemagglutinin Protein
No full textIdentification of pathotype of Avian Influenza Virus (AIV) subtype H5N1 isolates is very important. Thisresearch aimed to identify the pathotype of AIV subtype H5N1 isolated from household waterfowls in West Javabased on molecular markers of amino acid sequences of the Hemagglutinin (HA) cleavage site. Fragments of HAgenes of 21 isolates were amplified using RT-PCR with a primer pair that flanking the cleavage site region, andsequenced with dideoxy-termination method with ABI automatic sequencer (Applied Biosystems). Multiple alignmentof nucleotide and their deduced amino acid sequence were analyzed using ClustalW from MEGA 3.1 program.The result shows that all H5N1 isolates (21 isolates) possess polybasic cleavage sites with 2 patterns ofamino acid sequence, i.e QRERRRKKR (20 isolates) and QRESRRKKR (1 isolate). This finding indicates that all ofthe viruses isolated in this research were of highly pathogenic avian influenza (HPAI) strains.Keywords: cleavage site, waterfowls, HPA
Citrus reticulata's Peels Modulate Blood Cholesterol Profile and IncreaseBone Density of Ovariectomized Rats
No full textHormon Replacement Therapy is a common therapy for estrogen deficiency but in other side it will increase the risk of cardiovascular disease. Another alternative therapy which relatively more safe is using phytoestrogen. The Citrus reticulata’s peel contain flavanone and polimethoxyflavone which are suspected to give estrogenic effect, therefore it is potential to be used as phytoestrogen.The purpose of this study was to examine the estrogenic effect of Citrus reticulata’s peel extract in modulation of bone density and blood cholesterol profile of ovariectomized rats (OVX), an animal model of postmenopausal osteoporosis. Thirty six 7-weeks-old female Sprague Dawley rats were assigned to six groups: a SO group, an OVX group, an OVX+CMCNa group, an OVX+extract dose 500 mg/kgBW group, an OVX+extract dose 1000 mg/kgBW group, and an OVX+estradiol group. After 7 weeks, the rats were killed then blood and femoral were collected immediately. The rontgenogram indicated that extract and estradiol administration increase the bone density. And the data analysis with Oneway ANOVA test ,followed by Shceffé test (P 0.05) showed that extract can improve blood cholesterol profile in dose depend manner. These results suggest a possible role of Citrus reticulata’s peel extract as women’s health agent because of its beneficial effects on bone and lipids
Identification of Thymocyte Subset by Multicolor Flow Cytometry ED LSR II FACSDriva - FlowJo Software Analysis
No full textIn their development, thymocytes express different cell surface molecules that important for identification of thymocyte subset. It’s not easy to detect this cell surface molecules to determine the thymocyte subpopulation for research. Here we used multicolor flow cytometry ED LSR II FACSDriva - FlowJo software to identify of thymocyte subset from thymocyte sample solution using several antibodies such as mouse anti rat CD2-FITC, mouse anti rat CD45RC-PE, mouse anti rat CD4-APC, mouse anti rat CD8á-PerCP, mouse anti rat CD3-Biotin + PE-Cy7 or APC-Cy7. We determined double negative and single positive thymocyt subset (CD4 or CD8), found that the double negative thymocyte subset express CD2 and CD45RC. It was useful to determine the thymocyte subset using multicolor flow sitometry ED LSR II FACSDriva - FlowJo software
The Aquaeous Extract of Root Nodules Vigna radiata (rnVr) which Inoculated by Rhizobium as an Orally Available Anemia Therapeutic Candidate
No full textThe extract of root nodules Vigna radiata (rnVr) which inoculated by Rhizobium is considered beneficial as an orally available anemia therapeutic candidate, because it contain the leghemoglobin. The positive control mice (group I) were fed with the high nutrient pellet.The twelve mice (Mus musculus) was treated with the “taking rice pellet” that representing the low nutrient food for 21 days until they suffered anemia. Then, the anemia mice were treated orally with rnVr in different concentration groups:II. 0% III.33%; IV.67% and V.100%, respectively and fed with the “aking rice pellet”. After 14 days, the blood mice were collected from orbital sinus. The hemoglobin (Hb) concentration were analyzed by spectrophotometry and blood plasma profile protein were analyzed with electrophoresis (SDS-PAGE). All anemia mice that treated with rnVr showed the increasing of Hb and group that treated with 100% extract of rnVr could reach a normal Hb value, raising from 9.85 to 12.68 g/dL. There were observed the proteins which have molecule weight 36.5 and 35.7 kDa that indicated the existing erythropoietin. The increasing haemoglobin concentration and erythropoietin suggested if extract of rnVr could increasing red blood production and potential as an orally available anemia therapeutic candidate
Succession of Actinomycetes During Composting Proccess of Dairy-Farm Waste Investigated by Culture-Dependent and Independent Approaches
No full textMesophilic, thermophilic, and maturation phases were recognized in composting proccess. Temperaturechanges influence the microbial communities in compost within composting proccess. Actinomycetes account for alarger part of compost microbial population. The aim of this research was to study succession of actinomycetescommunity during composting of dairy-farm waste investigated by culture-dependent and independentapproaches.In culture-independent method, the succession of actinomycetes community was analyzed by nestedpolymerasechain reaction of ribosomal intergenic spacer (nested-PCR RISA) using spesific primer F243 and primerR23S followed by a second PCR using primers F968 and R23S. In culture-dependent method actinomycetes fromcompost were isolated on selective media, starch-nitrate medium and humic-acid + vitamins medium. DNA ofactinomycetes was extracted and amplified by repetitive sequence-based PCR (rep-PCR) using primer BOXA1R. Thebanding patterns were used to generate dendrograms by UPGMA clustering with NTSYS program. Microcosmcontaining sterile rice-straw and water which is inoculated with each actinomycetes isolates was used for examiningthe ability of each isolate in rice-straw degradation.The experiment results showed that succession of both bacteria and actinomycetes was occured withincomposting proccess of dairy-farm waste. Analysed by culture-independent method revealed that the highestcommunity of compost’s bacteria was on mesophilic, thermophilic, and maturation phases, respectively. WhereasPCR-nested RISA resulted the highest population of actinomycetes was on thermophilic, maturation, and mesophilicphases, respectively. By culture-dependent method was obtained 29 actinomycetes isolates from mesophilic phase,23 isolates from thermophilic phase, and 19 isolates from maturation phase. Genetic diversity analysis of the obtainedisolates showed the presence of phylogenetic grouping on each phase of composting proccess. This result illustratedthe occurance of succession of actinomycetes community in compost. The ability of each isolates in rice-strawdegradation was different, and SnT9 isolate was found to be a promising rice-straw degrader.Keywords: succession, actinomycetes, composting, nested-PCR RISA, rep-PC
Molecular Genotyping of HBV by using Nested PCR-RFLP among Hepatitis B Patients in Daerah Istimewa Yogyakarta Province and Surrounding Area
No full textHepatitis B virus (HBV) can be classified into 8 genotypes, genotype A to H. Genotype of HBV is important for clinical and etiological investigations. Research for HBV genotyping, HBV transmission study using nested PCR and HBV genotyping based on RFLP using restriction enzymes have been reported. However, both of those methods have been not applied for HBV genotyping study among hepatitis B patients in endemic area, like Indonesia yet. Molecular genotyping of HBV will describe epidemiology, pathogenesis and clinical implication of HBV. Combination of nested PCR and RFLP (nested PCR-RFLP) method to determine HBV genotype in Indonesia is still less information. The objectives of study were to develop a system for HBV genotyping by nested PCR combined with RFLP (nested PCR-RFLP) method based on nucleotide sequence of surface protein encodinggene (S gene) in HBV genome and to confirm HBV genotypes which predominantly found among hepatitis B patients in Daerah Istimewa Yogyakarta Province and surrounding area. Total of 149 sera from chronic hepatitis B patients from Daerah Istimewa Yogyakarta and surrounding areas were collected for in this work. Viral DNA were extracted from sera of hepatitis B patients and used as template for first round nested PCR amplification using outer primers set. Amplicons of first round PCR were used as template for second round amplification using inner primers set. Then, amplicons of second round nested PCR were restriction digested by Sty I and Bsr I enzymes. For HBV genotyping then the restriction products were analyzed by RFLP based on restriction pattern. Results showed that the first round nested PCR amplification generated DNA fragments of whole S gene in length 1.233 bp, and in second round nested PCR amplification using inner primer set generated DNA fragments 585 bp in length. Genotype analysis for all samples using nested PCR-RFLP methods by restriction digested of Sty I and Bsr I enzymes found only 2 HBV genotypes among hepatitis B patients, namely genotype B and C. Quantificationdata showed that most of hepatitis B patients found infected by HBV genotype B (92,8%), genotype C (3,6%) and unidentified genotype (3,6%). Nested PCR-RFLP methods for HBV genotyping is simple and inexpensive for clinical diagnostic in large scale