Antibodies titer of dogs immunized by anti-idiotypic vaccine detected by using enzyme linked immunosorbent assay

Rabies control programs, including extensive vaccination with attenuated or inactivated vaccines. However, such vaccines are not without problems and can have detrimental effects. Indeed attenuated vaccines can revert to a more virulent form, and inactivated vaccines may produce serious side effects. These facts, have led to the creation of a new generation of vaccines: recombinant-DNA vaccines, synthetic peptide vaccines, and anti-idiotypic vaccines. The aim of this study is to study the result of anti-idiotypic immunization methods in dogs detected by using enzyme linked immunosorbent assay (ELISA). Anti-idiotype antibodies against rabies (Ab2) were isolated from chicken blood, separated by means of ammonium sulfate precipitation, then dialyzed using PBS pH 8.0 for 24 hours at 2 – 8 oC and purified using affinity chromatography column. Three groups of dogs were immunized, group I was immunized intramuscularly (i.m) with purified IgY, group II was immunizded oraly (p.o) with purified IgY and group III was immunized intramuscularly (i.m) with rabies viral vaccines.  The antibody response (Ab3) was detected using Agar Gel Precipitation Test (AGPT). The efficacy of Ab3 was detected using ELISA. By ELISA, the result of immunization indicated that the level of Ab3 titers of anti-idiotypic vaccine immunized dogs intramuscularly are more than 0.5 IU/ml (protective according to WHO standard), and significantly higher than oraly immunization, but it significantly lower than Ab3 titers of rabies viral vaccine immunized dogs. The conclusion of this study is intramuscularly immunization of anti-idiotypic antibodies can induce protective immune response against rabies virus, although its lower than antibodies titer of viral vaccine, it has a good prospect for vaccine development in controlling rabie

Complementary monoclonal antibody-based dot ELISA for universal detection of H5 avian influenza virus

BACKGROUND: Rapid diagnosis and surveillance for H5 subtype viruses are critical for the control of H5N1 infection. RESULTS: In this study, H5 Dot ELISA, a rapid test for the detection of avian H5N1 influenza virus, was developed with two complementary H5 monoclonal antibodies. HA sequencing of escape mutants followed by epitope mapping revealed that the two Mabs target the epitope component (189(th )amino acid) on the HA protein but are specific for different amino acids (189Lys or 189Arg). Gene alignment indicated that these two amino acids are the most frequent types on this position among all of the H5 AIV reported in GeneBank. These two H5 Mabs were used together in a dot ELISA to detect H5 viral antigen. The detection limit of the developed test for multiple clades of H5N1 viruses, including clades 0, 1, 2.1, 2.2, 2.3, 4, 7, and 8, was less than 0.5 hemagglutinin units. The specificity of the optimized dot ELISA was examined by using 100 H5 strains, including H5N1 HPAI strains from multiple clades, 36 non-H5N1 viruses, and 4 influenza B viruses. No cross-reactivity was observed for any of the non-H5N1 viruses tested. Among 200 random poultry samples, the test gave 100% positive results for all of the twelve RT-PCR-positive samples. CONCLUSIONS: Considering that the test is convenient for field use, this H5 Dot ELISA can be used for on-site detection of H5N1 infection in clinical or environmental specimens and facilitate the investigation of H5N1 influenza outbreaks and surveillance in poultry

PEMBUATAN DAN STANDARISASI ANTIGEN AI H5N1 KOMERSIAL UNTUK MONITORING TITER ANTIBODI HASIL VAKSINASI AI DI INDUSTRI PETERNAKAN AYAM

Vaccination is one of the chosen strategy for controling AI H5N1 in Indonesia. Vaccination able to induce protective antibodies against AI but unable to inhibit viral infection. Determination of antibody titers in the serum from bird vaccinated with AI-H5N1 vaccine consisting of 2 or 3 different AI virus isolates difficult to be meassured if the antigen for HI test is uncalibrated yet. Furthermore, the determination of a minimum protective antibody titer against the challenge of AI virus circulating in the field at this time needs to be done.  This study aims to determine the H5N1 AI virus antigen for standart HI test and the minimum titre of antibodies that able neutralize virus infection. As much as 55 chickens were divided into 11 groups, 10 groups vaccinated with commercial AI vaccine and AI H5N1 field isolat antigen. Four types of commercial vaccines were veccinated to one group and seven other groups vaccinated with the antigen AI Legok 2004, Nagrak Ag 2009, Ag Lawang 2010, as well as polyvalent Ag combination of these three types of antigen. After third vaccinations, the presence of antibodieswere meassured by HI test. Serum with a titer test 26-28 were tested for the capability of virus neutralizationin using virus neutralization test against three different H5N1 AI virus field isolates. The test results showed that the H5N1 subtype AI virus antigen representative as standart antigen for HI test is antigen Legok 2004 and the minimum titer which able neutralize H5N1 AI virus field isolates 28

Effect of topical anti-Streptococcus mutans IgY gel on quantity of S. mutans on rats’ tooth surface

This study aims to evaluate the effect of anti-Streptococcus mutans IgY gel on quantity of S. mutans on rats’ tooth surface. Sprague Dawley rats were exposed intra-orally with S. mutans Xc and were fed a caries-inducing diet 2000. The 24 rats were divided into four groups: group A had their teeth coated with IgY gel; group B received sterilized water as a control; group C had their teeth coated with IgY gel starting on the 29th day; and group D had their teeth coated with a gel without IgY. Plaque samples were swabbed from the anterior teeth for S. mutans colony quantification, and saliva was collected to measure immunoreactivity by enzyme-linked immunosorbent assay. The results indicated that the quantity of S. mutans in rats treated with IgY gel showed significant difference compared with the controls. After coating with IgY anti-S. mutans gel, the mean immunoreactivity in rat saliva was higher than that of the no treatment group. In conclusion, topical application with anti-S. mutans IgY gel reduced the quantity of S. mutans on the tooth surface

Deteksi Spesies Brucella pada Kambing di Rumah Potong Hewan Jakarta

Brucellosis is a zoonosis and occupational diseases transmision. The diseases caused by bacterial and attack multiple species of animals. Common species that infects goats as the most pathogenic species (zoonotic) is Brucella melitensis; however, the species B. abortus could also infect goats. The study purposed to find out the brucellosis seropositive in goat in Jakarta slaughterhouse and to detect caused agent of brucellosis. Sampling was done through slaughtered goats that come from brucellosis endemic area. The samples were collected fromslaughtered mature female goats i.e serum, goat milk, vaginal swab, mamary gland, limphoglandula supramamary, limph, and uterus. The detection method was used i.e patological lession, serological, culture and PolymeraseChain Reaction (PCR) technique. The serological detection of brucellosis in goats was done parallelly between Rose Bengal Test (RBT), Complement Fixation Test (CFT) and Enzyme Linked Immunosorbent Assay (ELISA). The results of this study demonstrated that out of the 119 serum samples serologically tested, negative for RBT, one was positive for CFT and none were positive with ELISA. Patological observation in the Brucella predilection organs, there were 5 goat carcases showed pathological lession (vagina discharge, hemoragy at limphand limphoglandula, crumbly limph and there were pus in uterus). The serum samples that had reacted positively and the organs with pathological lesion were confirmed further with PCR, bacterial isolation and identification.The PCR test results and the culture of milk samples, vaginal swabs and organs did not reveal any Brucella spp bacteria (B. abortus, B. melitensis, B. ovis dan B. suis) and also vaccine strains of RB51. Based on these results, it was concluded that brucellosis in goats on Java Island was a 0.84% seropositive (confidence interval 95%; 0.00826 - 0.00854) (1/119), although the species of Brucella that had infected them remains unknown

Preparasi Imunoglobulin Yolk (IgY) Spesifik Virus Rabies untuk Pengembangan Kit Diagnostik

Penelitian ini bertujuan untuk memproduksi dan mengkarakterisasi IgY anti rabies sebagai bahan diagnostik. Ayam petelur usia produktif divaksinasi dengan vaksin rabies inaktif secara parenteral melalui rute intramuskular dengan dosis 0,5 ml sebanyak 2 kali. Keberadaan IgY pada telur dievaluasi dengan metode ELISA. Konsentrasi total protein IgY di hitung dengan metode Bradford. IgY dipurifikasi menggunakan dua metode yaitu : 1) pengendapan dengan NaCl, PEG 6000-amonium sulfat; 2) teknik Water Soluble Fraction (WSF), dilanjutkan pengendapan dengan PEG 6000-amonium sulfat.  Titer IgY spesifik  di tentukan dengan uji ELISA dan karakterisasi protein dengan metode SDS-PAGE. Hasil pengujian menunjukkan, antibodi anti-rabies dapat dideteksi pada kuning telur di minggu kedua setelah vaksinasi pertama. Purifikasi IgY dengan NaCl menghasilkan konsentrasi 331 µg/ml dan teknik WSF 184 µg/ml. Karakterisasi protein pada teknik NaCl menghasilkan 6 pita protein dengan berat molekul 164,16 kDa, 126,43 kDa, 97,36 kDa, 68,73 kDa, 40,76 kDa, 28,77 kDa sedangkan teknik WSF hanya terdiri dari 3 pita dengan berat molekul 94,03 kDa, 65,61 kDa, dan 31,94 kDa. Titer antibodi spesifik menggunakan teknik NaCl  lebih besar dari 0,5 IU/ml dan teknik WSF dengan titer antibodi di bawah 0,5 IU/ml . Berdasarkan hasil penelitian, dapat disimpulkan bahwa IgY spesifik rabies dapat diproduksi pada ayam petelur dan menghasilkan titer antibodi ≥ 0,5 IU/ml, dengan titer antibodi spesifik rabies sebesar ≥ 0,5 IU/ml

L3 Populations in Laying Hens Infected with 6,000 L2 of Ascaridia galli

The aim of the present study was to determine the survival of L3 populations in intestine ofchickens exposed to experimental Ascaridia galli infection. Nature female adult worm were obtained fromlumen of village chickens in a comercial abattoir in Bogor. The eggs (L1) obtained from uteri female adultworms were incubated in sterile aquadestilata at room temperature for 10-20 days developed embrionatedeggs (L2). Five groups (A-D) of 80 head chickens were infected with, 6000 L2 A. galli respectively. Thechickens of group A were infected six times with dose of each 1,000 L2 with an interval of one hour. Thechickens of group B were infected three times with dose of each 2,000 L2 with an interval of two hours.The chickens of group C were infected six times with dose of each 3,000 L2 with an interval of three hours. The chickens of group D were infected one time with single dose 6,000 L2. A. galli L3 were recovered from intestines of 80 heads chickens seven days after oesophagus inoculation with 6,000 L2.The result showed that total 702,000 L1 and 628,000 L2 collected from 124 A. galli female adult worms.The percentage of L1 developed L2 is 89.46% and L2 developed L3 is 11.27%. Significant survival of L3higher populations in intestine of chickens observed only in the group D. The results indicated thatchickens infected high dose of A. galli caused the decrease of host defence against ascaridiosis. Keywords: Ascaridia galli, embrionated eggs, larva

KONSENTRASI PROTEIN DAN PENENTUAN BERAT MOLEKUL EKSKRETORI/SEKRETORI L3 Ascaridia galli

Penelitian ini bertujuan menentukan konsentrasi dan berat molekul protein  ekskretori/sekretori larva (L3) Ascaridia galli (A. galli). Larva L3 diperoleh dari usus halus 100  ayam tujuh hari setelah pemberian dosis 6000 L2 melalui esofagus ayam. Sebanyak 5–10  L3 dikultur secara in vitro  dalam setiap ml medium Rosswell Park Memorial Institute (RPMI 1640), pH 6,8, tanpa merah fenol dalam inkubator pada temperatur 37 0C dan 5% CO2 selama 3 hari. Ke dalam medium ditambahkan 100 unit ml-1 penisilin G, 100 µg ml-1 streptomisin, 5 µg ml-1 gentamisin dan 0,25 µg ml-1 kanamisin. Ekskretori/sekretori dipreparasi dari produk metabolisme L3 yang dilepaskan ke dalam medium kultur. Untuk mendapatkan protein ekskretori/sekretori, medium kultur dipekatkan dengan vivaspin 30.000 MWCO, dan kuantitas protein dihitung dengan metode Bradford. Berat molekul protein ekskretori/sekretori divisualisasikan dengan sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS PAGE). Hasil penelitian  menunjukkan bahwa konsentrasi protein ekskretori/sekretori adalah 0,595 mg/ml dengan berat molekul 28 kDa