Professor of animal reproduction, Faculty of Veterinary Medicine, Bogor Agricultural University (IPB), Bogor - Indonesia. Director of Centre for Collaborative Research (CCR) Animal Biotechnology and Coral Reef Fisheries (ANBIOCORE), a center under the Sustainable Higher Education Research Alliance (SHERA) project, supported by USAID. Address: Bogor, Jawa Barat, Indonesia
Background: To obtain accurate measurements of cortisol (C) and testosterone (T) in Aceh cattle, ... more Background: To obtain accurate measurements of cortisol (C) and testosterone (T) in Aceh cattle, commercial enzyme-linked immunosorbent assay (ELISA) kits need to be carefully validated. Moreover, repeated freeze-thaw cycles during the storage of the samples may affect the stability of the hormones in the serum. Here, we test the reliability of C and T concentration measurements in the serum of Aceh cattle, obtained using commercial C and T ELISA kits designed to measure human C and T concentrations. Further, we evaluate the effect of repeated freeze-thaw cycles on the stability of C and T concentrations in the serum. Methods: Commercial C (Cat. no. EIA-1887) and T (Cat. no. EIA-1559) ELISA kits from DRG Instruments GmbH were validated through an analytical validation test (i.e., parallelism, accuracy, and precision) and a biological validation test (for C: effect of transportation on the C excretion; for T: the concentrations of T between bulls and cows). To test the effects of fre...
Background and Aim: Indonesia has two National Artificial Insemination centers and 17 Regional Ar... more Background and Aim: Indonesia has two National Artificial Insemination centers and 17 Regional Artificial Insemination Centers. The frozen semen production techniques differed between the centers, including the type of diluent and semen dilution technique. The aim of the research was to compare the quality of frozen Limousin bull semen diluted using different techniques. Materials and Methods: Semen was collected from three sexually mature Limousin bulls using an artificial vagina. Immediately after collection, the semen was evaluated macroscopically and microscopically. Semen that had >70% motile sperm and <20% sperm abnormality was divided into three tubes and diluted with skim milk-egg yolk (SMEY) using three different dilution techniques: One-step dilution (100% SMEY with 8% glycerol) at room temperature ([RT] 20°C until 25°C) two-step dilution (50% SMEY without glycerol at RT, stored at 5°C; and 50% SMEY with 16% glycerol after 1 h stored at 5°C); and three-step dilution ...
Livestock Embryo Center/Balai Embrio Ternak (BET) Cipelang is one of the government institution u... more Livestock Embryo Center/Balai Embrio Ternak (BET) Cipelang is one of the government institution under the supervision of Directorate General of Livestock and Animal Health Services, Ministry of Agriculture of the Republic of Indonesia. BET Cipelang has the main task of carrying out the production, development, and distribution of livestock embryo for all regions of Indonesia. BET Cipelang produced embryos using two methods, there are in vivo and in vitro. The produced embryos will be transferred to the recipient cow using embryo transfer technique. Embryo transfer biotechnology can be used to enhance genetic improvement and to increase marketing opportunities with purebred cattle. Because of their relatively low reproductive rate and long generation interval, embryo transfer is especially useful with cattle. The success of embryo transfer depends on factors associated with the embryo, the recipient or an interaction among the factors of the embryo and recipient [1]. ...
A biopsy of 1-5 blastomeres was taken from 86 bovine compacted morulae, using a technique, that d... more A biopsy of 1-5 blastomeres was taken from 86 bovine compacted morulae, using a technique, that did minimal damage to the zona pellucida. As soon as possible after the micromanipulation the embryos were frozen, without sealing the penetrated zona pellucida. In order to evaluate the viability of the biopsied frozen-thawed embryos, half of them were transferred to synchronized recipients while the remaining half were co-cultured to evaluate the developmental capacity. A control group of 43 intact embryos was subjected to the same procedures. This study revealed a slight, but not significant, decrease in the pregnancy rate of the biopsied embryos after freezing and thawing, although the embryos by co-culture with bovine oviduct epithelial cells revealed normal morphology and developmental rate. It is concluded that the described biopsy technique did not compromise the freezability of 7-day-old bovine embryos.
ContentsBovine demi-embryos were produced by micromanipulation of day 6–7 embryos from superovula... more ContentsBovine demi-embryos were produced by micromanipulation of day 6–7 embryos from superovulated cows. Monolayers of bovine oviduct epithelial cells (BOEC) were produced from slaughterhouse material. Half of the demi-embryos were cultured on a confluent layer of bovine oviduct epithelial cells either in Hams F-10 with 10% NCTC-135 and with 10% FCS or in Earles Medium 199 with 10% FCS for 4–24 hours, followed by freezing of the survivors, while the corresponding other halfs were used for sexing. three 16-Cell, 1 morula, 41 compact morulae, 4 early blastocysts, 7 blastocysts and 2 expanded blaszocysts were besected and cultured. An overall survival rate of 83% among the 63 in vitro cultured demi-embryos was seen.Bovine demi-embryos were produced by micromanipulation of day 6–7 embryos from superovulated cows. Monolayers of bovine oviduct epithelial cells (BOEC) were produced from slaughterhouse material. Half of the demi-embryos were cultured on a confluent layer of bovine oviduct epithelial cells either in Hams F-10 with 10% NCTC-135 and with 10% FCS or in Earles Medium 199 with 10% FCS for 4–24 hours, followed by freezing of the survivors, while the corresponding other halfs were used for sexing. three 16-Cell, 1 morula, 41 compact morulae, 4 early blastocysts, 7 blastocysts and 2 expanded blaszocysts were besected and cultured. An overall survival rate of 83% among the 63 in vitro cultured demi-embryos was seen.Inhalt: In vitro Kultivierung von geteilten Rinderembryonen mit bovinen Ovidukt-Epithel-AmmenzellenSechs bis sieben Tage alte Embryonen von superovulierten Rindern wurden mit Hilfe eines Mikromanipulators geteilt. Monolyer von bovinen Eileiterepithelzellen (BOEC) wurden aus Schlachthofmaterial gerüchtet. Eine Hälfte der Embryonen diente der Geschlechtsbestimmung, die andere Hälfte wurde auf konfluierenden Monolayern entweder mit Hams F-10, 10% NCTC-135 und 10′% FCS oder in TCM 199 rnit 10% FCS für 2 bis 4 Stunden kultiviert. Die überlebenden Stadien wurden eingefroren. Drei 16-Zeller, 1 Morula, 41 kompaktierte Morulae, vier frühe und sieben Blastozysten sowie 2 expandierte Blastocysten wurden geteilt und kultiviert. Insgesamt betrug die Oberlebensrate bei 63 in vitro kultivierten Zwillingsembryonen 83%.Sechs bis sieben Tage alte Embryonen von superovulierten Rindern wurden mit Hilfe eines Mikromanipulators geteilt. Monolyer von bovinen Eileiterepithelzellen (BOEC) wurden aus Schlachthofmaterial gerüchtet. Eine Hälfte der Embryonen diente der Geschlechtsbestimmung, die andere Hälfte wurde auf konfluierenden Monolayern entweder mit Hams F-10, 10% NCTC-135 und 10′% FCS oder in TCM 199 rnit 10% FCS für 2 bis 4 Stunden kultiviert. Die überlebenden Stadien wurden eingefroren. Drei 16-Zeller, 1 Morula, 41 kompaktierte Morulae, vier frühe und sieben Blastozysten sowie 2 expandierte Blastocysten wurden geteilt und kultiviert. Insgesamt betrug die Oberlebensrate bei 63 in vitro kultivierten Zwillingsembryonen 83%.
The structure of oocytes aspirated from the dominant and its subordinate follicles was investigat... more The structure of oocytes aspirated from the dominant and its subordinate follicles was investigated from the achievement of follicular dominance to ovulation. Ovulation was induced in 18 heifers and 5 cows by injection of cloprostenol at days 8-14 (day 0 = day of ovulation), and follicular development was monitored by ultrasonography. The animals were slaughtered at days 3-11, but animals slaughtered on days 8-11 were given a second injection of cloprostenol at day 7 to allow ovulation of the dominant follicle of the first follicular wave. Oocytes were aspirated from the dominant (largest) and two largest subordinate follicles and processed for transmission electron microscopy, whereas the follicular fluids were analyzed for concentrations of estradiol-17 beta (E2) and progesterone (P4). Dominant follicular growth was associated with increase in the concentration of E2 and P4 in the follicular fluid, which was E2-dominated. From days 3-7, the dominant oocytes had pronounced junctional contacts with the cumulus cells and a nonundulating nuclear envelope but showed an increase in the number of lipid droplets and a decrease in the size of Golgi complexes, the size of cortical granule clusters, and the number of microvilli stacks. After cloprostenol injection on day 7, but before the anticipated LH surge, the dominant oocytes showed a reduced oocyte-cumulus contact, vacuolization of the nucleolus, undulation of the nuclear envelope, and dispersal of the mitochondrial clusters. The morphological alterations occurring in the dominant oocytes before the anticipated LH surge are suggested to be a prerequisite for the oocyte to achieve the competence to undergo final maturation. Subordinate follicles ceased growing at about days 3-4 and their follicular fluid had low E2:P4 ratio or was P4-dominated. Subordinate oocytes displayed degenerative features in their cumulus investment and nuclear activation and maturation especially after day 5. The structural changes associated with oocyte degeneration showed similarities with the processes seen before and during final maturation of the dominant oocytes.
... Kusdiantoro Mohamad1,3, Mia Olsson2, Göran Andersson2,Bambang Purwantara1, Helena TA van Tol3... more ... Kusdiantoro Mohamad1,3, Mia Olsson2, Göran Andersson2,Bambang Purwantara1, Helena TA van Tol3, Sofia Mikko2, Heriberto Rodriguez-Martinez2, Ben ... Other cattle species (Lenstra and Bradley, 1999) are the yak (Bos grunniens) in and around Tibet, the gayal (Bos ...
Background: To obtain accurate measurements of cortisol (C) and testosterone (T) in Aceh cattle, ... more Background: To obtain accurate measurements of cortisol (C) and testosterone (T) in Aceh cattle, commercial enzyme-linked immunosorbent assay (ELISA) kits need to be carefully validated. Moreover, repeated freeze-thaw cycles during the storage of the samples may affect the stability of the hormones in the serum. Here, we test the reliability of C and T concentration measurements in the serum of Aceh cattle, obtained using commercial C and T ELISA kits designed to measure human C and T concentrations. Further, we evaluate the effect of repeated freeze-thaw cycles on the stability of C and T concentrations in the serum. Methods: Commercial C (Cat. no. EIA-1887) and T (Cat. no. EIA-1559) ELISA kits from DRG Instruments GmbH were validated through an analytical validation test (i.e., parallelism, accuracy, and precision) and a biological validation test (for C: effect of transportation on the C excretion; for T: the concentrations of T between bulls and cows). To test the effects of fre...
Background and Aim: Indonesia has two National Artificial Insemination centers and 17 Regional Ar... more Background and Aim: Indonesia has two National Artificial Insemination centers and 17 Regional Artificial Insemination Centers. The frozen semen production techniques differed between the centers, including the type of diluent and semen dilution technique. The aim of the research was to compare the quality of frozen Limousin bull semen diluted using different techniques. Materials and Methods: Semen was collected from three sexually mature Limousin bulls using an artificial vagina. Immediately after collection, the semen was evaluated macroscopically and microscopically. Semen that had >70% motile sperm and <20% sperm abnormality was divided into three tubes and diluted with skim milk-egg yolk (SMEY) using three different dilution techniques: One-step dilution (100% SMEY with 8% glycerol) at room temperature ([RT] 20°C until 25°C) two-step dilution (50% SMEY without glycerol at RT, stored at 5°C; and 50% SMEY with 16% glycerol after 1 h stored at 5°C); and three-step dilution ...
Livestock Embryo Center/Balai Embrio Ternak (BET) Cipelang is one of the government institution u... more Livestock Embryo Center/Balai Embrio Ternak (BET) Cipelang is one of the government institution under the supervision of Directorate General of Livestock and Animal Health Services, Ministry of Agriculture of the Republic of Indonesia. BET Cipelang has the main task of carrying out the production, development, and distribution of livestock embryo for all regions of Indonesia. BET Cipelang produced embryos using two methods, there are in vivo and in vitro. The produced embryos will be transferred to the recipient cow using embryo transfer technique. Embryo transfer biotechnology can be used to enhance genetic improvement and to increase marketing opportunities with purebred cattle. Because of their relatively low reproductive rate and long generation interval, embryo transfer is especially useful with cattle. The success of embryo transfer depends on factors associated with the embryo, the recipient or an interaction among the factors of the embryo and recipient [1]. ...
A biopsy of 1-5 blastomeres was taken from 86 bovine compacted morulae, using a technique, that d... more A biopsy of 1-5 blastomeres was taken from 86 bovine compacted morulae, using a technique, that did minimal damage to the zona pellucida. As soon as possible after the micromanipulation the embryos were frozen, without sealing the penetrated zona pellucida. In order to evaluate the viability of the biopsied frozen-thawed embryos, half of them were transferred to synchronized recipients while the remaining half were co-cultured to evaluate the developmental capacity. A control group of 43 intact embryos was subjected to the same procedures. This study revealed a slight, but not significant, decrease in the pregnancy rate of the biopsied embryos after freezing and thawing, although the embryos by co-culture with bovine oviduct epithelial cells revealed normal morphology and developmental rate. It is concluded that the described biopsy technique did not compromise the freezability of 7-day-old bovine embryos.
ContentsBovine demi-embryos were produced by micromanipulation of day 6–7 embryos from superovula... more ContentsBovine demi-embryos were produced by micromanipulation of day 6–7 embryos from superovulated cows. Monolayers of bovine oviduct epithelial cells (BOEC) were produced from slaughterhouse material. Half of the demi-embryos were cultured on a confluent layer of bovine oviduct epithelial cells either in Hams F-10 with 10% NCTC-135 and with 10% FCS or in Earles Medium 199 with 10% FCS for 4–24 hours, followed by freezing of the survivors, while the corresponding other halfs were used for sexing. three 16-Cell, 1 morula, 41 compact morulae, 4 early blastocysts, 7 blastocysts and 2 expanded blaszocysts were besected and cultured. An overall survival rate of 83% among the 63 in vitro cultured demi-embryos was seen.Bovine demi-embryos were produced by micromanipulation of day 6–7 embryos from superovulated cows. Monolayers of bovine oviduct epithelial cells (BOEC) were produced from slaughterhouse material. Half of the demi-embryos were cultured on a confluent layer of bovine oviduct epithelial cells either in Hams F-10 with 10% NCTC-135 and with 10% FCS or in Earles Medium 199 with 10% FCS for 4–24 hours, followed by freezing of the survivors, while the corresponding other halfs were used for sexing. three 16-Cell, 1 morula, 41 compact morulae, 4 early blastocysts, 7 blastocysts and 2 expanded blaszocysts were besected and cultured. An overall survival rate of 83% among the 63 in vitro cultured demi-embryos was seen.Inhalt: In vitro Kultivierung von geteilten Rinderembryonen mit bovinen Ovidukt-Epithel-AmmenzellenSechs bis sieben Tage alte Embryonen von superovulierten Rindern wurden mit Hilfe eines Mikromanipulators geteilt. Monolyer von bovinen Eileiterepithelzellen (BOEC) wurden aus Schlachthofmaterial gerüchtet. Eine Hälfte der Embryonen diente der Geschlechtsbestimmung, die andere Hälfte wurde auf konfluierenden Monolayern entweder mit Hams F-10, 10% NCTC-135 und 10′% FCS oder in TCM 199 rnit 10% FCS für 2 bis 4 Stunden kultiviert. Die überlebenden Stadien wurden eingefroren. Drei 16-Zeller, 1 Morula, 41 kompaktierte Morulae, vier frühe und sieben Blastozysten sowie 2 expandierte Blastocysten wurden geteilt und kultiviert. Insgesamt betrug die Oberlebensrate bei 63 in vitro kultivierten Zwillingsembryonen 83%.Sechs bis sieben Tage alte Embryonen von superovulierten Rindern wurden mit Hilfe eines Mikromanipulators geteilt. Monolyer von bovinen Eileiterepithelzellen (BOEC) wurden aus Schlachthofmaterial gerüchtet. Eine Hälfte der Embryonen diente der Geschlechtsbestimmung, die andere Hälfte wurde auf konfluierenden Monolayern entweder mit Hams F-10, 10% NCTC-135 und 10′% FCS oder in TCM 199 rnit 10% FCS für 2 bis 4 Stunden kultiviert. Die überlebenden Stadien wurden eingefroren. Drei 16-Zeller, 1 Morula, 41 kompaktierte Morulae, vier frühe und sieben Blastozysten sowie 2 expandierte Blastocysten wurden geteilt und kultiviert. Insgesamt betrug die Oberlebensrate bei 63 in vitro kultivierten Zwillingsembryonen 83%.
The structure of oocytes aspirated from the dominant and its subordinate follicles was investigat... more The structure of oocytes aspirated from the dominant and its subordinate follicles was investigated from the achievement of follicular dominance to ovulation. Ovulation was induced in 18 heifers and 5 cows by injection of cloprostenol at days 8-14 (day 0 = day of ovulation), and follicular development was monitored by ultrasonography. The animals were slaughtered at days 3-11, but animals slaughtered on days 8-11 were given a second injection of cloprostenol at day 7 to allow ovulation of the dominant follicle of the first follicular wave. Oocytes were aspirated from the dominant (largest) and two largest subordinate follicles and processed for transmission electron microscopy, whereas the follicular fluids were analyzed for concentrations of estradiol-17 beta (E2) and progesterone (P4). Dominant follicular growth was associated with increase in the concentration of E2 and P4 in the follicular fluid, which was E2-dominated. From days 3-7, the dominant oocytes had pronounced junctional contacts with the cumulus cells and a nonundulating nuclear envelope but showed an increase in the number of lipid droplets and a decrease in the size of Golgi complexes, the size of cortical granule clusters, and the number of microvilli stacks. After cloprostenol injection on day 7, but before the anticipated LH surge, the dominant oocytes showed a reduced oocyte-cumulus contact, vacuolization of the nucleolus, undulation of the nuclear envelope, and dispersal of the mitochondrial clusters. The morphological alterations occurring in the dominant oocytes before the anticipated LH surge are suggested to be a prerequisite for the oocyte to achieve the competence to undergo final maturation. Subordinate follicles ceased growing at about days 3-4 and their follicular fluid had low E2:P4 ratio or was P4-dominated. Subordinate oocytes displayed degenerative features in their cumulus investment and nuclear activation and maturation especially after day 5. The structural changes associated with oocyte degeneration showed similarities with the processes seen before and during final maturation of the dominant oocytes.
... Kusdiantoro Mohamad1,3, Mia Olsson2, Göran Andersson2,Bambang Purwantara1, Helena TA van Tol3... more ... Kusdiantoro Mohamad1,3, Mia Olsson2, Göran Andersson2,Bambang Purwantara1, Helena TA van Tol3, Sofia Mikko2, Heriberto Rodriguez-Martinez2, Ben ... Other cattle species (Lenstra and Bradley, 1999) are the yak (Bos grunniens) in and around Tibet, the gayal (Bos ...
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