Simple and ultra-fast recognition and quantitation of compounded monoclonal antibodies: Application to flow injection analysis combined to UV spectroscopy and matching method.

Compounding of monoclonal antibody (mAbs) constantly increases in hospital. Quality control (QC) of the compounded mAbs based on quantification and identification is required to prevent potential errors and fast method is needed to manage outpatient chemotherapy administration. A simple and ultra-fast (less than 30 s) method using flow injection analysis associated to least square matching method issued from the analyzer software was performed and evaluated for the routine hospital QC of three compounded mAbs: bevacizumab, infliximab and rituximab. The method was evaluated through qualitative and quantitative parameters. Preliminary analysis of the UV absorption and second derivative spectra of the mAbs allowed us to adapt analytical conditions according to the therapeutic range of the mAbs. In terms of quantitative QC, linearity, accuracy and precision were assessed as specified in ICH guidelines. Very satisfactory recovery was achieved and the RSD (%) of the intermediate precision were less than 1.1%. Qualitative analytical parameters were also evaluated in terms of specificity, sensitivity and global precision through a matrix of confusion. Results showed to be concentration and mAbs dependant and excellent (100%) specificity and sensitivity were reached within specific concentration range. Finally, routine application on "real life" samples (n = 209) from different batch of the three mAbs complied with the specifications of the quality control i.e. excellent identification (100%) and ±â€¯15% of targeting concentration belonging to the calibration range. The successful use of the combination of second derivative spectroscopy and partial least square matching method demonstrated the interest of FIA for the ultra-fast QC of mAbs after compounding using matching method.

Group C streptococcal psoas abscess associated with a homolateral hip joint prosthesis infection: a case report.

An abscess in the psoas muscle is rare and frequently misdiagnosed. A delay in the diagnosis can increase its mortality rate. Some clinical signs can help the clinician but they all are not always present, and not at the same time. We describe in this paper a case report of an association between a psoas abscess and a homolateral hip joint prosthesis infection. It was suspected because of no improvement in clinical state despite treatment of the abscess by antibiotics and drainage, and it required finally other complementary therapeutic solutions. The pathogenic microorganism was a group C streptococcus. We discuss all these points and thereafter we suggest some recommendations for the clinician.

In vitro protein-binding characteristics of delavirdine and its N-dealkylated metabolite.

This study was performed to determine delavirdine protein-binding characteristics as well as those of its N-dealkylated metabolite (N-DLV). Initial studies of 36 microM delavirdine and 30 microM N-DLV in solutions of plasma, albumin 4 g%, alpha-1-acid glycoprotein (AAG) 100 mg% or immune globulin (IVIG) 5 g% were conducted. Delavirdine (12, 36 and 73 microM) and N-DLV (10, 30 and 60 microM) were then studied alone and in combination in plasma and various concentrations of albumin. Studies were done in triplicate using equilibrium dialysis. The mean delavirdine fraction unbound (fu) in plasma, albumin, IVIG and AAG was 0.013, 0.033, 0.752 and 0.912 while the mean fu of N-DLV in these same protein solutions was 0.139, 0.195, 0.329 and 0.359. In plasma and albumin, a greater fu was observed at higher delavirdine concentrations and no significant changes in fu were noted with the addition of N-DLV. An increase in delavirdine fu was noted as the albumin concentrations decreased. The fu of N-DLV increased significantly as the concentration of albumin decreased as well as with decreasing N-DLV concentration. The potential implications of extensive delavirdine binding to plasma proteins, primarily albumin, are discussed.

Radiological method for preoperative determination of isometric attachment points of an anterior cruciate ligament graft.

The purpose of this study was to develop a radiological method which would be preoperatively available to help determine the best graft placement (with respect to isometricity as well as absence of graft impingement) for all knees. The radiological method is described in full detail. We also present the most significant experimental work supporting our development. Firstly, we studied the path followed by radio-opaque objects inserted in the mobile tibia around the fixed femur. Secondly, we compared the distances measured between selected femoral and tibial points radiologically (according to our method) and clinically (with a currently available isometer). The main results were: (1) every tibial point considered moves on an arc of a circle centered or a corresponding femoral point. We should then speak of pairs of isometric points instead of a single femoral isometric zone; (2) the more posterior the tibial point, the more anterior and distal the corresponding femoral point and vice versa; (3) the distance variations induced by rotation did not exceed 1.5 to 2.5 mm when measured either radiologically or clinically; (4) on the radiological and clinical measurements, the difference of length variations during flexion was also very small (mean 0.22 mm; SD 1.2 mm). We conclude that this very simple method allows us to find the femoral transition line for every knee (whatever its size, shape or dynamics). It aids preoperative planning in anterior cruciate ligament graft reconstruction.

Frequency of early in utero HIV-1 infection: a blind DNA polymerase chain reaction study on 100 fetal thymuses.

OBJECTIVE: To estimate the prevalence of in utero transmission of HIV-1 through the second trimester. MATERIAL AND METHODS: One hundred consecutive, unselected, intact fetuses, beyond 15 weeks gestational age (mean, 22.4 weeks) were studied. These were obtained following spontaneous intrauterine deaths (n = 4), miscarriages (n = 4), and elective mid-trimester terminations (n = 92), eight of which were fetuses with malformations from HIV-1-positive pregnancies. Coded DNA extracts from the fetal thymuses were tested blindly by polymerase chain reaction in three laboratories using a total of six different primer pairs. RESULTS: Two thymuses tested positive [95% confidence interval (Cl), 0.2-7]. Results from the three laboratories were consistent in all 100 cases. The two fetuses with HIV in the thymus both tested positive in other organs, demonstrating systemic HIV infection. The first fetus, whose mother had advanced AIDS, had died in utero and had diffuse toxoplasmosis. The second died following extremely premature delivery in a pregnancy complicated by repeated bleeding. HIV infection was observed in none of the 92 fetuses that resulted from elective mid-trimester terminations (95% Cl, 0-4). CONCLUSION: The frequency of early in utero HIV infection appears to be low, compared with transmission rates in infants born to HIV-1-infected mothers, suggesting that transmission occurs mostly later in pregnancy and/or at delivery. Specific risk factors may have implications in the occurrence of early as opposed to late transmission.

Characterization of dual HIV-1 and HIV-2 serological profiles by polymerase chain reaction.

OBJECTIVE: To evaluate the frequency of dual HIV-1 and HIV-2 DNA sequences in patients with dual serological profiles. DESIGN: We tested 40 samples from AIDS patients living in Abidjan, Côte d'Ivoire. METHODS: Dual serological reactivity was determined by double Western blot and two enzyme-linked immunosorbent assays with recombinant proteins and synthetic peptides as antigens. The Western blot was considered to show dual reactivity when sera reacted with at least two glycoproteins and one core protein of each virus. HIV DNA sequences were detected by hybridization to radiolabelled probes of polymerase chain reaction (PCR) products amplified using specific primers. RESULTS: Both HIV-1 and HIV-2 DNA sequences were detected in four out of 11 samples with a dual serological profile and in four out of 24 samples with anti-HIV-1 antibodies only. CONCLUSION: These results show that dual HIV-1 and HIV-2 serological profiles are not always due to infection by both viruses, and emphasize the need for a combination of serological and PCR assays for the appraisal of these viral infections.

Vaccine protection of chimpanzees against challenge with HIV-1-infected peripheral blood mononuclear cells.

Because human immunodeficiency virus (HIV) can be transmitted as cell-free virus or as infected cells (cell-associated virus), vaccines must protect against infection by both viral forms. Vaccine-mediated protection of nonhuman primates against low doses of cell-free HIV-1, HIV-2, or simian immunodeficiency virus (SIV) has been demonstrated. It is now shown that multiple immunizations of chimpanzees with HIV-1 antigens protected against infection with cell-associated virus. Protection can persist for extended periods (one animal had not been exposed to viral antigens for 1 year before challenge). These results show that it is possible to elicit long-lasting protective immunity against cell-associated HIV-1.

[The treatment of rigid hallux using Swanson's silastic implant (single and double stem). Clinical, radiological and podobarographic review with a 16-year maximum follow-up]. / Traitement de l'hallux rigidus par les implants en Silastic de Swanson (single et double stem). Revue clinique, radiologique et podobarographique avec 16 ans de recul maximal.

Twenty-eight cases of hallux rigidus treated with the Swanson Silastic implant (single and double stem) were reviewed, with an 8-year follow-up. The skin complication rate was significant. Long-term patient satisfaction was good. Radiologic findings were alarming: implants seem to wear out quickly on the articular side, and granulomatous reactions develop around the stems. Dynamic pedobarography shows decreased pressure under the first ray, with transfer of the weight to the midmetatarsal heads, sometimes on the external edge of the foot. The authors review the international literature on the etiology, pathology and treatment of hallux rigidus. Surgical indications and techniques are specified.

Immunization of chimpanzees confers protection against challenge with human immunodeficiency virus.

Sustained high titers of neutralizing antibodies were elicited in three chimpanzees after sequential injections of different human immunodeficiency virus 1 (HIV-1) antigen preparations derived from the HIV-1 BRU strain that included whole inactivated virus or purified recombinant proteins and then synthetic peptides identical to the major HIV-1 neutralizing epitope V3. The animals were challenged i.v. with 40 chimpanzee infectious doses (equivalent to 100 tissue culture 50% infectious doses) of a stock of HIV-1 IIIB isolate. After 6 mo of follow-up, all three animals appeared uninfected by serologic and virologic criteria, including polymerase chain reaction analysis and failure to isolate virus from peripheral blood lymphocytes, bone marrow, and lymph node tissue. Of two chimpanzees monitored for 1 yr, virus was isolated initially from one animal at 32 weeks, but the second chimpanzee was virus negative by all assays through 12 mo; the third animal has remained virus negative through 9 mo of follow-up. These results indicate that it is possible to elicit protection against, or significantly delay infection of, HIV-1 by immunization, thus laying the foundation for development of an HIV-1 vaccine.

Active suppression of host-vs-graft reaction in pregnant mice. VII. Spontaneous abortion of allogeneic CBA/J x DBA/2 fetuses in the uterus of CBA/J mice correlates with deficient non-T suppressor cell activity.

The mammalian fetus has been viewed as an unusually successful type of "allograft" and "unexplained" spontaneous abortion as a possible example of maternal rejection. Previous studies have shown the presence of small lymphocytic suppressor cells in the murine decidua which block the generation and reactivation of anti-paternal cytotoxic T lymphocytes (CTL) and lymphokine-activated killer cells (LAK) by elaborating a factor that inhibits the response to interleukin 2 (IL 2). A deficiency of these suppressor cells was associated with implants of xenogeneic Mus caroli embryos in the Mus musculus uterus which are infiltrated by maternal lymphoid cells and aborted. We have also shown a deficiency of such suppressor cells in the lymph nodes draining the uterus of CBA/J females in the process of aborting their semi-allogeneic CBA X DBA/2 F1 progeny. CBA/J females possess significantly lower levels of decidua-associated non-T suppressor cells on day 8.5 to 10.5 of allopregnancy than do mothers that will produce large litters of live babies. The F1 embryos are infiltrated by maternal lymphocytes prior to abortion, and the infiltration and abortion rate appears to be augmented by pre-immunization with paternal DBA/2 spleen cells. Susceptibility to spontaneous abortion is dependent upon maternal age and strain of male mate, and the high abortion rate of CBA/J mated to DBA/2 males can be reduced by immunization with BALB/c spleen cells. The CBA/J x DBA/2J mating combination provides a model of spontaneous abortion in which immunologic factors play an important role and demonstrates that the association between deficiency of decidua-associated suppressor cells and xenopregnancy failure also holds true for the failure of allopregnancies resulting from natural within-species mating.

Immunoregulatory molecules of trophoblast and decidual suppressor cell origin at the maternofetal interface.

The mammalian fetus expresses a variety of paternal histocompatible, oncofetal, and trophoblast antigens against which the mother can mount an immune response. Survival of the "fetal graft" appears to depend upon local immunosuppressive mechanisms in lymph nodes draining the uterus and at the intrauterine implanation site itself. Nonspecific not-T-Fc-receptor-bearing small lymphocytes containing cytoplasmic granules present in successfully allopregnant mice can suppress both the generation of maternal-antipaternal killer T cells and the infiltration of cytotoxic T lymphocytes into sponge-matrix allografts during the effector phase of the immune response. These suppressor cells are deficient at the implantation sites of xenogeneic and allogeneic mouse embryos that are susceptible to maternal immunity and are destined to resorb. A soluble suppressor factor of approximately 100,000 daltons in size can be obtained from the suppressor cells and acts to block the response of T cells to interleukin-2 by interfering with IL-2 receptors. The development of the suppressor cells in the decidua requires certain hormonal signals as well as signals provided by trophoblast cells. Freshly explanted or cultured murine trophoblast cell lines elaborate soluble factor(s) that are active in recruitment or activation of suppressor cells. Since suppressor cells may be isolated from decidua of successfully allopregnant humans, the suppressor cell mechanism and its regulation may represent a key factor in the protection of the "fetal allograft" from rejection by maternal immunity.

Strain-dependence and cellular aspects of the acceleration of age-dependent shift in class-specific helper and suppressor activity in the thymus of MRL/Mp mice by the LPR gene.

Previous studies of the immunoregulatory activity of thymocytes from SJL/J mice have shown loss of suppressor activity for the antibody response by 24 weeks of age with appearance of helper activity. At the same time, suppressor cells developed which inhibit the generation of cytotoxic T lymphocytes (CTL). We now show a similar pattern of helper and suppressor activity in MRL/Mp mice. Presence of the lpr/lpr genotype significantly accelerated the onset of these changes in thymocyte activity. A similar pattern of thymocyte activity was not detected in C57B1/6 mice. In aged MRL-lpr mice, evidence of increased suppressor cell activity for the CTL response could be demonstrated in spleen, and the suppressor was sensitive to treatment with anti-thy 1.2 + complement. The magnitude of the deficiency in the CTL response in MRL-lpr mice was greater than could be accounted for by suppressor cell activity alone. Measurement of the frequency of CTL precursors (CTLP), the yield of CTL per CTLP, and the ability to produce and to respond to interleukin 2 (IL-2) indicated that a drop in CTLP frequency, subnormal generation of IL-2, and probably an intrinsic defect in the responsiveness of MRL-lpr CTLP to IL-2 was contributing to the defective CTL response. We were not able to link suppressor T cells with reduced responsiveness to IL-2. Ageing involves different patterns of change in immunoregulatory T-cell subsets in different strains of mice, depending on their genetic constitution. The general implications of this conclusion for prediction of immune dysfunction with age in genetically distinct members of an outbred population are discussed.

Active suppression of host-vs-graft reaction in pregnant mice. VI. Soluble suppressor activity obtained from decidua of allopregnant mice blocks the response to IL 2.

The mammalian fetus expresses a variety of antigens against which the maternal immune system can react and which in an allogeneic mating bears paternal transplantation antigens. Although these antigens may be expressed on the fetal trophoblast cells that contact maternal uterine decidua, the "fetal allograft" is not usually rejected. Previous studies have demonstrated the presence of nonspecific non-thymus-derived suppressor cells in the lymph nodes draining the uterus and in decidua of laboratory mice undergoing first allogeneic pregnancy. These suppressor cells appeared to be small lymphocyte cells that inhibit the generation of cytotoxic T lymphocytes (CTL) in vitro and in vivo and elaborate a nonspecific non-MHC-restricted soluble suppressor activity when cultured for 48 hours at 37 degrees C in vitro. We now report that soluble suppressor activity obtained from the decidua (DS) of allopregnant C3H/HeJ mice inhibits both the primary and secondary (memory) CTL response in vitro but does not inhibit lysis of target cells by preformed CTL. DS did not suppress the proliferation of YAC lymphoma cells, P-815 cells, or a C3H placental trophoblastoma line. Suppressor activity was obtained from anti-thy-1.2 + complement-resistant cells in the decidua, could also be obtained from the decidua of allopregnant CD1 nu/nu mice, and was associated with a single peak of activity of approximately 100,000 daltons on Sephacryl 200 chromatography. Suppression could not be overcome by adding either crude or HPLC-purified IL 2 to the mixed lymphocyte cultures in vitro, and both crude and column-purified suppressor factor inhibited the IL 2-dependent proliferation of H-Y cells (a cloned T cell line with NK activity). Furthermore, DS inhibited the IL 2-dependent generation of cytotoxic effector cells in vitro in the absence of allogeneic stimulator cells. Thus, a soluble suppressor factor obtained from non-T cells present in the decidua of successfully allopregnant mice could block the response to IL 2 and inhibit the generation of both specific and nonspecific cytotoxic effector cells. The significance of this inhibition with respect to survival of the "fetal allograft" is discussed.

Suppressor cells in human decidua.

Decidual suppressor cells have been found in the murine system. These cells are absent at the implantation sites of interspecies mouse embryos which become infiltrated by maternal cytotoxic cells. Suppression is also deficient at the sites of the spontaneous resorption of fetuses in allomated intraspecies pregnancies. This study was carried out to determine whether similar suppressor cells were present in the decidua during successful human allopregnancies. Decidua was obtained from gestations of 13 to 15 weeks and from term gestations, and the lymphocytes were tested for their ability to suppress the response of their peripheral blood lymphocytes to concanavalin A. Eight of eight 13- to 15-week decidual lymphocytes proved to be suppressive. At term seven of twelve lymphocyte preparations at a lower concentration of cells and six of seven at a higher concentration showed suppression. Suppressor cells appear to be present in human decidua and may play a role in preventing maternal immunologic attack on the allogenic embryo, thereby preventing spontaneous abortion.

[Gingivitis and recession in the child (100 cases)]. / Les gingivites et les récessions chez l'enfant (à propos de 100 cas).

An epidemiological study was made on 100 children 5 to 18 years old, from the Paris region, who showed precocious gingival pathology. Then, the various factors, liable to prevent such defects before the point of no return is reached, were investigated. They are hygiene and its type and frequency. The responsibility lies with the dental profession, educators and public dental health services.