Estudos sôbre a excitação química da córtex cerebral (Ação da acetilcolina)

The author has studied the influence of acetylcholine solutions directly applied on the motor cortex of dogs, cats monkeys and rabbits. For this purpose small squares of filter paper were soaked in the acetylcholine solution and soon afterwards laid on the motor cortex. Solutions varying from 0,2 to 10 per cent have been experimented. It has been shown that local application of the solutions on the motor points, previously localized by induction coil, produced motor reactions. It has been found, in the dogs that 10 per cent acetylcholine solutions cause localized muscular twitchings (clonus) in almost all the animals experimented. Generalised epileptiform convulsions were obtained in44,4% of the dogs. Convulsions were also obtained by employing 1 per cent solution of acetylcholine. Definite response has been obtained with 0,2 per cent solution. Failure of motor action, pointed out by other authors, has been related to the use of anesthetics. Convulsions were easily produced by rapid light mechanical stimulations of the skin covering the muscles in conection with the excited motor point, and the application on the motor point of acetylcholine. The results on monkeys can be summarized as follows. Two species of monkeys were experimented: Cebus capucinus and Macaca mulata. In the monkeys C. capucinus generalised convulsive reactions were induced with actylcholine solutions in a concentration as low as 0,5 per cent. Motor reaction or convulsive seizeres were obtained in seven of the eight monkeys used. Three monkeys M. mulata were stimulated with 10 per cent acetylcholine solution but only localized muscular contraction hae been observed. Similar results has been obtained on the motor cortex of cats and rabbits. One of the three cats employed has shown epileptiform convulsions and the remaining only localized muscular contractions. In the rabbits muscular twitchings have been also induced. The sensitizing power of eserine on the action of acetylcholine has been also searched. The results indicate that a previous application of eserine solution on the motor center, potentiates the action of acetylcholine. The intensity of the muscular twitchings is greater than the obtained before the application of the eserine solution. Generalised epileptiform convulsions sometimes appeared following the use of lower concentrations of acetylcholine than those previously employed. Experiments have been carried out by injecting eserine and prostigmine by parenteral route. A dosis dufficient for induce small muscular tremors did not enhance obviously the motor effects produced by the application of the acetylcholine solutions on the motor cortex. From seven dogs experimented, all previously tested for convulsive seiruzes by application of 1 and 10 per cent acetylcholine solution with negative results, only one has shown epileptiform convulsions after the injection of prostigmine. Morphine has also been tested as facilitating substance for convulsions induced by acetylcholine. Six from the nine dogs submitted to the experiments, developed epileptiform seizures after injection of morphine and stimulation of the motor cortex with acetylcholine. (Table IV). In another series of experiments atropine and nicotine have been studied as for to their action on the motor effects of acetylcholine. Nicotine has a strong convulsant action, even when employed in very high concentration. Since a depressant effect has not appeared even by the applications of high concentrations of nicotine in the motor corteõ of dogs, unlike the classical observations for the autonomus nervous system, it was not possible to verify the action of acetylcholine on a motor center paralised by nicotine. It is important to not that the motor phenomena observed after the first aplication of acetylcholine, can desappear by the renewal of the pieces of filter paper soaked in the acetylcholine solution. Atropine, either applied on the motor point in low concentration, or injected in sufficient amount for inhibiting the “muscarinic effects” of acetylcholine on the autonomous nervous system, did not prevent the motor reactions of acetylcholine on the cerebral cortex.

Infecção do macaco sul americano "saguí" (Callitrix jacchus, Linneu 1758) com o Plasmodium Knowlesi

Foram observadas infecções no macaco americano "Sagui" (Callitrix jacchus, Linneu 1758), quando inoculados com Plasmodium knowlesi. Inoculações massiças intra-venosas, são mortais em cerca de 10 dias. Com inoculações intra-musculares de menores quantidades de parasitos, foram verificados alguns casos de curas espontaneas. As características morfológicas e o seu conhecido ciclo de 24 horas taes como observados na infecção do macaco Rhesus, permanecem as mesmas no macaco estudado. O vigor do "sagui" em cativeiro é nitidamente inferior ao do "Rhesus" mas como seu preço de custo é trinta vezes menor, deve ser êle considerado como mais um animal útil para as pesquisas em malaria experimental.

Subsídio ao melhor conhecimento do tifo exantemático neotrópico no Brasil: presença do virus no leite

In articles, already published, we have proved that the strain V. B. of Brazilian virus, goes through the placenta (Macacus rhesus) (1) and the apparently normal gastro-intestinal tube (1934-1937) (Canis familiaris) (2). Today we present the idea that the Brazilian virus can reach the milk of an animal even when the latter has only the unapparent disease. In former articles (**), we have shown that the goat (Capra hircus) can be an excellent reservoir of Brazilian virus, having the strain V. B. in its blood and presenting a Weil Felix reaction high and in “group”, with the disease unapparent. When the goats are bred in the laboratory, and even in some foci of the disease, they give a negative Weil Felix, being zero for all the nine strains of Proteus. In the interior of Brazil, in many localities, goats substitute cows, in supplying milk for children and adults, and in some districts goat’s milk is considered superior to cow’s milk, possessing marvellous qualities for men, women an children. Having proved, now, that goat’s milk can contain the virus even when the animal presents nothing clinically, and having also shown that this virus goes through the digestive tube apparently sound, it is easy to understand how infants-in-arms, that is, only a few months old, living in strictly domestic surroundings, can contract the disease; we have many such cases on record. Protocol of the experiments: Goat nº 2, white, January 1948. This animal had been inoculated with the V. B. strain of the Brazilian virus in June 1947, via intra-peritoneal, presenting nothing then, not even a feverish reaction. On that occasion it was not possible to isolate the virus of the blood, although the Weil Felix reaction was positive, high and in “group”. Now January 17, 1948, seven months later, the same animal was reinoculated with a semple of virus V. B. in the same manner (intra-peritoneal) two days after bringing forth two sturdy kids. The virus V. B. was obtained from guinea-pig n. 7170 whose thermic graph was as follows: Temperatura – 38,8 – 39,1 – 39,5 – 39,4 –39,8 – 40,4 – 40,2 – 40,1 - + Necropsy – Typical lesions. The spleen weighed 5 grammes. With 3c.c. of emulsion from the nervous system of this guinea-pig, we inoculated not only the goat, as also two guineapigs, number 14 and number 5. The following is the thermic graph of one: - Guinea-pig n. 14 – 38,9 – 39,1 – 39,2 – 39.2 – 40,7 – 41,0 – 40,5 – 40,4 – 40,1 - + Typical lesions. Guinea-pig n. 2 presented the following thermic graph after the infective inoculation: - 39,5 – 39,7 – 39,7 – 39,7 – 39,5 – 39,3 – 39,5 – 39,5 – 39,5 – etc. Clinically, this animal presented nothing unusual, feeding well and suckling the kids normally. The Weil Felix reaction was positive, in “group” high very similar to the reaction obtained in June 1947, with the first infective inoculation. On the third, fourth, fifth, sixth and seventh day after the infective inoculation, we took milk from the goat and inoculated male guinea-pigs via intra-celular and via intra-peritoneal, giving 5 c.c. to each animal. Guinea-pig n. 4663, inoculated with 5 c.c. of milk, via intra-muscular, taken on the third day of the infectaive inoculation, presented the following thermic graph: - 38.8 (*) – 39,1 – 39,0 – 39,1 – 40,1 – 40,1 – 40,8 (**) – 40,8 – Killed – Typical deisions (***). The virus V. B. of this goat, circulated naturally in the blood up to the third day, having passed into the milk, producing nothing in the kids, on account of the natural resistance of these animals to the disease. The Weil Felix reaction and that of Widal for the Burcellas suis, abortus and militensis were negative for the goat and the kids. It is remarkable that, even with inoculation of the living virus after a period of seven months we cannot get a real and absolute immunity of sensitive animals. We shall return to this subject later. The hart Mazama simplicicornis may be a carrier of the virus in Brasil. The experimental serum against the virus of Exanthematic neotropical typhus has not protected guinea-pigs.

Miocardite no macaco Cebus após inoculações repetidas com Schizotrypanum cruzi

Transmission of Chagas disease is realized through contamination of ocular conjunctiva, mucosa or skin with infected dejections eliminated by the insect vectors of Schizotrypanum cruzi (Triatoma infestans, Panstrongylus megistus and Rhodnius prolixus). The triatomid bugs live in holes and craks in the walls, in beds, behind trunks, etc. Found in primitive mud huts covered with thatched roofs, and so the human dwellers have many chances to contract the disease, reinfections being reasonably more to expect than a single inoculation. Experimental work reproducing those natural conditions is welcomed as some important features in the pathologic picture of the disease such as the extensive myocardial fibrosis seen in chronic cases are still incompletely known. Microscopic changes were studied in the heart muscle of seven Cebus monkeys infected by S. cruzi. This animal survives the acute stage of the disease and so is particularly suited to experiments of long duration in which several inoculations of S. cruzi are performed. Three different strains of S. cruzi isolated from acute cases of Chagas' disease were employed. One monkey was injected in the skin with infected blood and necropsied after 252 days. Two monkeys were three times, and one, eight times infected in skin, one of them with contaminated blood, and two with contaminated blood and dejections from infected bugs. The necropsies were performed after 35, 95 and 149 days. One monkey was three times inoculated through the intact ocular conjunctiva (one time with infected blood, two times with dejections from infected bugs), and one time through the wounded buccal mucosa, and necropsied after 134 days. Another monkey was six times inoculated, four times through the intact ocular conjunctiva (one time with contaminated blood, three times with dejections from infected bugs) and two times injected in the skin with infected blood, and necropsied after 157 days. Finally, another monkey was nine times inoculated, four times through the intact ocular conjunctiva (one time with infected blood, and three times with dejections from infected bugs), and five times injected in the skin (four times with contaminated blood, and one time with dejections from infected bugs), and necropsied after 233 days. The microscopic picture was uniform presenting, however, considerable individual variations, and was represented by diffuse interstitial myocarditis, frequently more (marked in the right ventricle base of the heart), accompanied by lymphatic stasis. The infiltration consists of macrophages, plasma cells and lymphocytes, the cellular reaction having sometimes a perivascular distribution, involving the auriculo-ventricular system of conduction, endocardium, epicardium and cardiac sympathetic gangliae. The loss of cardiac muscle fibers was always minimal. Leishmanial forms of S. cruzi in myocardial fibers are scanty and, in two cases, absent. Fatty necrosis in the epicardium was noted in two cases. Obliterative changes of medium-sized branches of coronary arteries (hypersensitivity reaction?) and multiple infarcts of the myocardium was found in one instance. The diffuse myocarditis induced by S. cruzi in several species of monkeys of the genus Cebus observed after 233 days (several inoculations) and 252 days (single inoculation) is not associated with disseminated fibrosis such as is reported in chronic cases of Chagas' disease. Definite capacity of reversion is another characteristic of the interstitial myocarditis observed in the series of Cebus monkeys here studied. The impression was gained that repeated inoculation with S. cruzi may influence the myocardial changes differently according to the period between the reinoculations. A short period after the first inoculation is followed by more marked changes, while long periods are accompanied by slight changes, which suggests an active immunisation produced by the first inoculation. More data are required, however before a definite statement is made on this subject considering that individual variations, the natural capacity of reversion of the interstitial myocarditis and the employement of more than a species of Cebus monkeys probably exerts influence also in the results here reported.

Toxoplasmose em primatas não humanos: II - Tentativas de infecções experimentais em Macacca mulatta, Cebus apella e Callithrix jacchus; e pesquisa de anticorpos em várias espécies de platyrrhinus

Após uma revisão da toxoplasmose experimental em primatas não humanos, são relatadas as tentativas, sem êxito, para provocar toxoplasmose aguda e fatal em dois rhesus (Macacca mulatta), um infante e outro jovem, por inoculação e reinoculação de uma amostra humana, usando diferentes vias e doses maciças e ainda com a ministração de decametasona. Do mesmo modo, não teve sucesso a tentativa para induzir a doença fatal em um Cebus apella adulto, pela via peritoneal. Porém a toxoplasmose-infecção nesses 3 animais, foi comprovada pela elevação da temperatura (39 a 41ºC), pela positividade da reação de Sabin-Feldman (1:64 - 1:256) e pelo isolamento de toxoplasmas em camundongos inoculados com material do Cebus. Por outro lado, em um Callithrix jacchus pela inoculação peritoneal, foi provocada doença grave e fatal com focos necróticos e abundãncia de toxoplasmas no baço e fígado, e isolamento dos parasitas em camundongo. De 54 símios do Nõvo Mundo, submetidos a RSF, todos foram negativos, com exceção de um Saimiri que se mostrou positivo a 1:16 (18%). Uma análise do problema Toxoplasmose-Primatas não humanos, com o apoio na revisão da literatura e nas nossas próprias observações (ver também o trabalho anterior) permite as seguintes conclusões: em seu habitat natural os primatas não humanos não são expostos ao Toxoplasma. Isso deve estar relacionado aos hábitos arborícolas e à sua alimentação vegetariana e insetívora; b) os casos descritos de toxoplasmose natural nos símios se referem a animais de cativeiro; e, mesmo nestas condições, é excepcional a infecção espontânea dos catarrinos; c) os catarrinos apresentam, além disso uma grande resistência à indução da toxoplasmose experimental, a qual não é devida à presença de anticorpos circulantes. Essa resistência parece não ser rompida pela administração de corticosteróides; embora às vêzes o seja pela inoculação de doses maciças de toxoplasmas, e geralmente nos animais jovens; d) essa resistência é menor nos platirrinos e parece não existir nos platirrinos inferiores e nos Prosimii.

Panencefalite subaguda esclerosante: estudo comparativo entre as lesões humanas e as experimentais determinadas por agente encefalitogênico de origem humana

Neste trabalho estudamos as alterações histopatológicas encontradas no sistema nervoso central de dois pacientes com "panencefalite subaguda esclerosante" comparando-as com as modificações estruturais determinadas no sistema nervoso central de sete macacos rhesus nos quais este material foi inoculado. Os animais apresentaram sinais de comprometimento neurológico, traduzido por caquexia e paralisia do trem posterior, após um longo período de incubação, em torno de 18 meses. Dois animais morreram antes de qualquer manifestação neurológica, de infecção pulmonar intercorrente acidental. Nas passagens sucessivas houve um encurtamento do periódo de incubação para cerca de 40 dias. As alterações histopatológicas encontradas, consistiram, nos casos humanos, em leptomeningite focal, focos de neuronofagia, granulomas corticais e nos núcleos basais, grande perda da população neuronal com ocasional estado esponjoso do córtice cerebral, infiltrados perivasculares, e gliose da substância branca, sem perda de mielina. No material experimental foram observadas estas mesmas modificações, se bem que de caráter muito menos intenso. Tanto no material humano como no experimental a mielina estava praticamente normal. Sugere-se que o quadro anátomo-clínico chamado "panencefalite subaguda esclerosante (SSPE) possa ser determinado, não apenas pelo vírus do sarampo, mas também por outros vírus, especialmente os do grupo papova, já encontrado por outros autores, em casos de "panencefalite subaguda esclerosante".

Development of natural products as drugs acting on central nervous system

We have recenty studied several natural product constituents which have effects on the CNS. (1) Tetrahydropalmatine (THP) and its analogues were isolated from Corydalis ambigua and various species of Stephania. (+)-THP and (-)-THP posses not only analgesic activity, but also exert sedative-tranquillizing and hypnotic actions. Results of receptor binding assay and their pre-and post-synaptic effects on dopaminergic system indicate that (-)-THP and (-)-stepholidine are dopamine receptor antagonists while (+)-THP is a selective dopamine depletor. (2) 3-Acetylaconitine (AAC) is an alkaloid isolated from Aconitum flavum. The relative potency of analgesic action of AAC was 5.1-35.6 and 1250-3912 times that of morphine and aspirin, respectively. The analgesic effect of AAC was antagonized by naloxone, but was eliminated by reserpine. In monkeys, after AAC was injected for 92 days, no abstinence syndrome was seen after sudden AAC withdrawal or when challenged with nalorphine. (3) Huperzine A (Hup-A) is an alkaloid isolated from Huperzia serrata which was found to be a selective ChE inhibitor and could improve learning and retrieval process. Preliminary clinical studies showed that Hup-A improve short-and long-term memory in patients of cerebral arteriosclerosis with memory impairment. (4) Ranamargarin is a new tetradecapeptide isolated from the skin of the Chines frog Rana margaratae. This peptide may mainly act on NK-1 receptor.

Simian malaria in Brazil

In Brazil simian malaria is widely spread, being frequent in the Amazon region (10% of primates infected) and even more in the forested coastal mountains of the Southeastern and Southern regions (35% and 18% infected, respectively), but absent in the semi-arid Northeast. Only two species of plasmoidia have been found: the quartan-like Plasmodium brasilianum and the tertian-like P. simium, but the possible presence of other species is not excluded. P. brasilianum is found in all enzootic foci, but P. simium was detected only on the coast of the Southeastern and Southern regions, between parallels 20-S and 30-S. Nearly all hosts are monkeys (family Cebidae, 28 species harbouring plasmodia out of 46 examined) and very rarely marmosets or tamarins (family Callitrichidae, I especies out of 16). P. brasilianum was present in all infected species, P. simium in only two. The natural vector in the Southeastern and Southern regions was found to be Anopheles cruzi, but has not been conclusively identified in the Amazon. One natural, accidental human infection due to P. simium was observed. There is no evidence of the relation of the simian to human malaria in the Southeastern and Southern regions, where human malaria was eradicated in spite of the high rates of monkeys infected, but in the Amazon recent serological studies by other workers, revealing high positivity for P. brasilianum/P. malariae antibodies in local indians, would suggest that among them malaria might be regarded as a zoonosis.

Immunogenicity and antigenicity of the N-term repeat amino acid sequence of the Plasmodium falciparum P126 antigen

The P126 protein, a parasitosphorus vacuole antigen of Plasmodium falciparum has beenshoen to induce protective immunity in Saimiri and Aotus monkeys. In the present work we investigated its immunogenicity. Our results suggest that the N-term of P126 is poorly immunogenic and antibody response against the P126 could be under a MHC restricted control in C57BL/6(H-2b) mice, which could be problematic in ternms of a use of the P126 in a vaccine program. However, we observed that a synthetic peptide, copying the 6 octapeptide repeat corresponding to the N-term of the P126, induces an antibody response to the native molecule in C57BL/6 non-responder mice. Moreover, the vaccine-P126 recombinant induced anmtibodies against the N-term of the molecule in rabbits while the unprocessed P126 did not.

A recombinant hybrid protein as antigen for an anti-blood stage malaria vaccine: a study on the conservation of a protective component

Recently we have shown that two hybrid proteins expressed in Escherichia coli confer protective immunity to Aotus monkeys against an experimental Plasmodium falciparum infection (Knapp et al., 1992). Both hybrid proteins carry a sequence containing amino acids 631 to 764 of the serine stretch protein SERP (Knapp et al., 1989b). We have studied the diversity of this SERP region in field isolates of P. falciparum. Genomic DNA was extracted from the blood of six donors from different endemic areas of Brazil and West Africa. The SERP region encoding amino acids 630 to 781 was amplified by polymerase chain reaction (PCR) and sequenced. Only conserved amino acid substitutions in maximally two positions of the analyzed SERP fragment could be detected which supports the suitability of this SERP region as a component of anti-blood stage malaria vaccine.

A study on the pathogenesis of human cerebral malaria and cerebral babesiosis

Cerebral complications are important, but poorly understood pathological features of infections caused by some species of Plasmodium and Babesia. Patients dying from P. falciparum were classified as cerebral or non-cerebral cases according to the cerebral malaria coma scale. Light microscopy revealed that cerebral microvessels of cerebral malaria patients were field with a mixture of parazited and unparazited erythrocytes, with 94% of the vessels showing parasitized red blood cell (PRBC) sequestration. Some degree of PRBC sequestration was also found in non-cerebral malaria patients, but the percentage of microvessls with sequestered PRBC was only 13% Electron microscopy demonstrated knobs on the membrane of PRBC that formed focal junctions with the capillary endothelium. A number of host cell molecules such as CD36, thrombospondim (TSP) and intracellular adhesion molecule I (ICAM-1) may function as endothelial cell surfacereports for P. falciparum-infected erythrocytes. Affinity labeling of CD36 and TSP to the PRBC surface showed these molecules specifically bind to the knobs. Babesia bovis infected erythrocytes procedure projections of the erythrocyte membrane that are similar to knobs. When brain tissue from B. bovis-infected cattle was examined, cerebral capillaries were packed with PRBC. Infected erythrocytes formed focal attachments with cerebral endothelial cells at the site of these knob-like projections. These findings indicate that cerebral pathology caused by B. bovis is similar to human cerebral malaria. A search for cytoadherence proteins in the endothelial cells may lead to a better understanding of the pathogenisis of cerebral babesiosis.

Mechanisms of protective immunity against asexual blood stages of Plasmodium falciparum in the experimental host Saimiri

In the Saimiri monkey, an experimental host for human malaria, acquired protection against Plasmodium falciparum blood stages depends on the IgG antibody populations developed. In vivo protective anti-falciparum activity of IgG antibodies is correlated with the in vivo opsonizing activity promoting phagocytosis of parasited red bloood cells. In contrast, non protective antibodies inhibit this mechanism by competing at the target level. A similar phenomenon can be and human infection. Anti-cytoadherent and anti-rosette antibodies developed by Saimiri and humans prevent the development of physiopathological events like cerebral malaria which can also occur in this experimental host. Furthermore, transfer to protective human anti-falciparum IgG antibodies into infected Saimiri monkeys exerts an anti parasite activity as efficient as that observed when it is transfered into acute falciparum malaria patients, making the Saimiri an even more attractive host. Studies on the role of immunocompetent cells in the protective immune reponse are still in their infancy, however the existance of a restricted polymorphism of MHC II class molecules in the Saimiri confers additional theoretical and practical importance to this model.

Development of sporogonic cycle of Plasmodium vivax in experimentally infected Anopheles albimanus mosquitoes

The sporogonic cycle of Plasmodium vivax was established and maintained under laboratory conditions in two different strains of Anopheles albimanus mosquitoes using as a parasite source blood from human patients or from Aotus monkeys infected with the VCC-2 P.vivax colombian isolate. Both the Tecojate strain isolate from Guatemala and the Cartagena strain from the colombian Pacific coast were susceptible to infections with P.vivax. A higher percentage of Cartagena mosquitoes was infected per trial, however the Tecojate strain developed higher sporozoite loads. Intravenous inoculation of Aotus monkeys with sporozoites obtained from both anopheline strains resulted in successful blood infections. Animals infected with sporozoites from the Tecojate strain presented a patent period of 21-32 days whereas parasitemia appeared between days 19-53 in monkeys infected with sporozites from Cartagena strain.

Activity of 9-acridanone-hydrazone drugs detected at the pre-postural phase, in the experimental schistosomiasis mansoni

The compound Ro-15.5458/000, derivative in the class of 9-acridanone-hydrazones, was found to be effective against Schistosoma mansoni in mice, killing almost all the skin schistosomules (24 hr after infection), when administered at the dose of 100 mg/kg. In experiments carried out with Cebus monkeys, the drug was shown to be fully effective at 25 mg/kg, 7 days after infection. These data, associated with the good results obtained earlier at the post-postural phase of schistosomiasis, allow the inference that this promising compound may be important in the set of antischistosomal drugs, depending on further toxicological and clinical tests.

CHO expression of a novel human recombinant IgG1 anti-RhD antibody isolated by phage display.

Replacement of the hyperimmune anti-Rhesus (Rh) D immunoglobulin, currently used to prevent haemolytic disease of the newborn, by fully recombinant human anti-RhD antibodies would solve the current logistic problems associated with supply and demand. The combination of phage display repertoire cloning with precise selection procedures enables isolation of specific genes that can then be inserted into mammalian expression systems allowing production of large quantities of recombinant human proteins. With the aim of selecting high-affinity anti-RhD antibodies, two human Fab libraries were constructed from a hyperimmune donor. Use of a new phage panning procedure involving bromelin-treated red blood cells enabled the isolation of two high-affinity Fab-expressing phage clones. LD-6-3 and LD-6-33, specific for RhD. These showed a novel reaction pattern by recognizing the D variants D(III), D(IVa), D(IVb), D(Va), D(VI) types I and II. D(VII), Rh33 and DFR. Full-length immunoglobulin molecules were constructed by cloning the variable regions into expression vectors containing genomic DNA encoding the immunoglobulin constant regions. We describe the first, stable, suspension growth-adapted Chinese hamster ovary (CHO) cell line producing a high affinity recombinant human IgG1 anti-RhD antibody adapted to pilot-scale production. Evaluation of the Fc region of this recombinant antibody by either chemiluminescence or antibody-dependent cell cytotoxicity (ADCC) assays demonstrated macrophage activation and lysis of red blood cells by human lymphocytes. A consistent source of recombinant human anti-RhD immunoglobulin produced by CHO cells is expected to meet the stringent safety and regulatory requirements for prophylactic application.