Free Antibody Research Articles

DR5-targeted, chemotherapeutic drug-loaded nanoparticles induce apoptosis and tumor regression in pancreatic cancer in vivo models

Pancreatic cancer is usually advanced and drug resistant at diagnosis. A potential therapeutic approach outlined here uses nanoparticle (NP)-based drug carriers, which have unique properties that enhance intra-tumor drug exposure and reduce systemic toxicity of encapsulated drugs. Here we report that patients whose pancreatic cancers express elevated levels of Death Receptor 5 (DR5) and its downstream regulators/effectors FLIP, Caspase-8, and FADD had particularly poor prognoses. To take advantage of elevated expression of this pathway, we designed drug-loaded NPs with a surface-conjugated αDR5 antibody (AMG 655). Binding and clustering of the DR5 is a prerequisite for efficient apoptosis initiation, and the αDR5-NPs were indeed found to activate apoptosis in multiple pancreatic cancer models, whereas the free antibody did not. The extent of apoptosis induced by αDR5-NPs was enhanced by down-regulating FLIP, a key modulator of death receptor-mediated activation of caspase-8. Moreover, the DNA topoisomerase-1 inhibitor camptothecin (CPT) down-regulated FLIP in pancreatic cancer models and enhanced apoptosis induced by αDR5-NPs. CPT-loaded αDR5-NPs significantly increased apoptosis and decreased cell viability in vitro in a caspase-8- and FADD-dependent manner consistent with their expected mechanism-of-action. Importantly, CPT-loaded αDR5-NPs markedly reduced tumor growth rates in vivo in established pancreatic tumor models, inducing regressions in one model. These proof-of-concept studies indicate that αDR5-NPs loaded with agents that downregulate or inhibit FLIP are promising candidate agents for the treatment of pancreatic cancer.

Annals for Educators - 2 June 2020

Annals for Educators - 2 June 2020

Evaluate the fPSA for Discriminate between Prostate Cancer Patients (PCa) and Benign Prostatic Hyperplasia (BPH) Depending on PSA Fucosylation by Using Nano-Lectin Immunoassay

Background: The objective was to evaluate the fPSA for discriminate between prostate cancer patients (PCa) and benign prostatic hyperplasia (BPH) depend on the variations in PSA fucosylation (FucI±1-6/3GlcNAc carbohydrates) by using Aleuria aurantia lectin (AAL) . Materials and Methods: The new method lectin-immunoassays were established for the investigation of PSA fucosylation by measuring the fluorescence of Eu(III)-chelate-labeled nanoparticles by microplate flourometer. To the identification of FucI±1-6/3GlcNAc carbohydrates on PSA surface , the lectin-immunoassays using free PSA specific monoclonal antibody as acapture and AAL coupled with Eu(III)-chelatelabeled nanoparticles using as probe for detecting fPSA . these methods compared with conventional immunoassay ELISA Result and discussion: The nanoparticle-based lectin-immunoassay method we showed a statistical significant (p < 0.001) in fPSA level depend on the fucosylation from PCa patients compared to healthy control, and also clear significant (p < 0.001) when comparing HBP patients to PCa patients but HBP compared with healthy control there is non-significant the (p=0.086) in the nanoparticle-based lectinimmunoassay methods. while in the immunoasssy ELISA was showed deffrent result .

Capture-Layer Lateral Flow Immunoassay: A New Platform Validated in the Detection and Quantification of Dengue NS1.

The lateral flow immunoassay (LFIA) is the most successful point-of-care testing (POCT) method to date. In the case of clinical biomarkers that require quantification, it remains a challenge to quantitate those biomarkers using the lateral flow immunoassay remains a challenge due to the cost of the reader and possibly the type of marker used. In the present work, a new concept of a platform LFIA device configuration is proposed in which different, aligned membrane components, some already existing in the classical lateral flow immunoassay, and the others created with special new functions in the present device. As the sample containing the target analyte passes through the aforementioned membranes, the target analyte will initially interact with a target-specific antibody-conjugated to horseradish peroxidase (HRP). Thereafter, the newly formed immunocomplex will diffuse through a proprietary capture membrane (that ensures that the nontarget-bound antibodies do not continue further and thus remain “captured” to that specific area). This is done by having the target molecules (or components thereof) immobilized onto the said capture layer. The target-bound immunocomplexes will then be allowed by the system configuration to continue further to the last layer, where the signal will be generated and quantified. Thus, in the absence of the target analyte in the sample, the free antibodies will be filtered at the capture layer by preimmobilized analyte molecules, thus preventing a false positive signal to occur. We validated the concept in the detection of dengue NS1 protein in view of making a triage test. The sample containing NS1 will first meet HRP-conjugated NS1-specific antibodies and become attached, thus producing an NS1-specific antibody–HRP immunocomplex. The sample then flows through the blocking layer, where the immunocomplex is unchallenged and thus allowed to reach the last “absorbent” pad, incorporating the substrate for the HRP marker. In the case of a positive test, a signal is generated, that is proportional to the amount of immunocomplexes (and therefore the NS1 concentration), and then analyzed and measured at the absorbent pad. Any unbound anti-NS1 antibody will be stopped at the blocking matrix by preimmobilized NS1, so there will be no false positive. As this study is the initial study of a novel configuration, much of the work comprised of optimization steps, such as determining the required NS1 membrane-immobilization concentration and the required target-specific capture antibody concentration. Our immunoassay was tested with spiked buffer and serum samples to mimic the clinical conditions, with a range of NS1 concentrations, and was found, at this time, to be fivefold more sensitive than a gold standard enzyme-linked immunosorbent assay (ELISA) test (5 ng mL–1) performed in our laboratory. This method shows another form of LFIA that has the potential to be quantitative (at least semiquantitative), albeit not solving the reader cost; however, unlike the regular LFIA, we do not use nanobeads but instead enzymes, allowing, in theory, greater sensitivity, while retaining the one-step procedure. The test is accurate and has low production costs.

Autoantibodies targeting apolipoprotein A-I are associated with cardiovascular disease outcomes

Abstract Cardiovascular disease (CVD) is characterized by an inflammatory immune response associated with disease progression. Apolipoprotein A-I (ApoA-I), the major protein of high-density lipoprotein (HDL), has been identified as the target of antibodies in patients, but their role in disease progression is poorly understood. Recently, we identified immune complexes composed of ApoA-I and IgG (ApoA-I/IgG ICs) in human plasma, which are associated with delayed disease progression. Studies in a large prospective cohort of cardiovascular disease are currently underway. The mechanistic impact of these observations and the dichotomy between free and bound antibodies urge continued exploration of the anti-ApoA-I antibody response in mouse models of inflammation. To complement our efforts with plasma from human subjects, we evaluated anti-ApoA-I antibody induction in mice in response to immune stimuli including western diet, immunization with ApoA-I epitopes, induction of hypercholesterolemia, or combinations of all three. Anti-ApoA-I antibody profiles and subclass composition were characterized by ELISA and compared between groups. Most notably, mice exhibit elevated levels of free antibody responses toward ApoA-I with age, irrespective of gender or diet composition. These data indicate that autoantibodies targeting ApoA-I are naturally occurring in both mice and humans and prompt continued investigation into their role in cardiovascular disease progression.

Inhibition of Wnt signaling by Frizzled7 antibody-coated nanoshells sensitizes triple-negative breast cancer cells to the autophagy regulator chloroquine.

Despite improvements in our understanding of the biology behind triple-negative breast cancer (TNBC), it remains a devastating disease due to lack of an effective targeted therapy. Inhibiting Wnt signaling is a promising strategy to combat TNBC because Wnt signaling drives TNBC progression, chemoresistance, and stemness. However, Wnt inhibition can lead to upregulation of autophagy, which confers therapeutic resistance. This provides an opportunity for combination therapy, as autophagy inhibitors applied concurrently with Wnt inhibitors could increase treatment efficacy. Here, we applied the autophagy inhibitor chloroquine (CQ) to TNBC cells in combination with Frizzled7 antibody-coated nanoshells (FZD7-NS) that suppress Wnt signaling by blocking Wnt ligand/FZD7 receptor interactions, and evaluated this dual treatment in vitro. We found that FZD7-NS can inhibit Axin2 and CyclinD1, two targets of canonical Wnt signaling, and increase the expression of LC3, an autophagy marker. When FZD7-NS and CQ are applied together, they reduce the expression of several stemness genes in TNBC cells, leading to inhibition of TNBC cell migration and self-renewal. Notably, co-delivery of FZD7-NS and CQ is more effective than either therapy alone or the combination of CQ with free FZD7 antibodies. This demonstrates that the nanocarrier design is important to its therapeutic utility. Overall, these findings indicate that combined regulation of Wnt signaling and autophagy by FZD7-NS and CQ is a promising strategy to combat TNBC.

Plasmodium falciparum Merozoite Associated Armadillo Protein (PfMAAP) Is Apically Localized in Free Merozoites and Antibodies Are Associated With Reduced Risk of Malaria.

Understanding the functional role of proteins expressed by Plasmodium falciparum is an important step toward unlocking potential targets for the development of therapeutic or diagnostic interventions. The armadillo (ARM) repeat protein superfamily is associated with varied functions across the eukaryotes. Therefore, it is important to understand the role of members of this protein family in Plasmodium biology. The Plasmodium falciparum armadillo repeats only (PfARO; Pf3D7_0414900) and P. falciparum merozoite organizing proteins (PfMOP; Pf3D7_0917000) are armadillo-repeat containing proteins previously characterized in P. falciparum. Here, we describe the characterization of another ARM repeat-containing protein in P. falciparum, which we have named the P. falciparum Merozoites-Associated Armadillo repeats protein (PfMAAP). Antibodies raised to three different synthetic peptides of PfMAAP show apical staining of free merozoites and those within the mature infected schizont. We also demonstrate that the antibodies raised to the PfMAAP peptides inhibited invasion of erythrocytes by merozoites from different parasite isolates. In addition, naturally acquired human antibodies to the N- and C- termini of PfMAAP are associated with a reduced risk of malaria in a prospective cohort analysis.

Abstract B01: Nanoparticle multispecific T-cell engagers for the treatment of multiple myeloma

Abstract Introduction: Multiple myeloma (MM) is the second most prevalent hematologic malignancy. Chimeric antigen receptor (CAR)-T cells and bispecific T-cell engagers targeting BCMA, CS1, and CD38 are promising approaches for MM. However, using these targets individually limits the efficacy of these immunotherapy options due to the variable expression levels of these markers, leading to inefficient eradication of MM. Studies have attempted to develop CAR-T cells and T-cell engagers with multispecific targeting; however, these are still technically very challenging and are highly expensive. In this study, we developed nanoparticles with monovalent or multivalent specificity against MM targets and engage T cells, by chemical conjugation of monoclonal antibodies against each molecule to the surface of the liposome. Bi-specific nanoparticles with anti CD38/CD3, BCMA/CD3 or CS1/CD3 were termed nanoBiTEs, while the multispecific particles with anti BCMA/CS1/CD38/CD3 were termed nanoMuTEs. Methods: Liposomes were prepared using the thin-film hydration method followed by extrusion. nanoBiTEs and nanoMuTEs were developed by chemical conjugation of monoclonal antibodies against BCMA, CS1, and/or CD38; together with anti CD3 onto the liposomes. We have tested the expression of BCMA, CS1, and CD38 antigens in MM cell lines and primary MM patient samples, and we analyzed the binding of the CD38/CD3, BCMA/CD3, and CS1/CD3 nanoBiTEs, as well as the BCMA/CS1/CD38/CD3 nanoMuTEs to these cells by flow cytometry and confocal microscopy. Binding specificity was assessed by blocking the target with corresponding free antibodies. CD25 expression, a T-cell activation marker, was assessed for CD8 and CD4 T cells when incubated with or without nanoBiTEs and nanoMuTEs. Moreover, we tested the efficacy of the nanoBiTEs and nanoMuTEs against MM cells in vitro using the 3D tissue engineered bone marrow (3DTEBM) myeloma model. Finally, we tested the efficacy of the nanoBiTEs and nanoMuTE against MM tumors in vivo in xenograft MM models. Results: The expression of BCMA, CS1 and CD38 was variable in MM cell lines and patient samples, and so was the binding of each nanoBiTEs, while the binding of the nanoMuTE was higher and more universal than each nanoBiTE. Similar patterns were observed in activation of CD8 T cells (no significant activation was observed in CD4 T cells) and in eradication of MM cell lines and primary patient samples in vitro. In vivo, the nanoBiTEs extended the survival and reduced the tumor burden in the MM bearing mice compared to untreated and T cell alone groups, and nanoMuTE experiments are still ongoing. Conclusions: The nanoBiTEs and nanoMuTEs were bound specifically to MM cells and initiated CD8 T-cell activation that resulted in MM eradication in vitro and in vivo, while the effect of the nanoMuTEs was more universal and efficacious than each nanoBiTE. These nanoparticles are an inexpensive method of engaging T cells for the treatment of MM and represent a new class of immunotherapy that can be expanded to other tumors. Citation Format: Kinan Alhallak, Jennifer Sun, Barbara Muz, Christopher Egbulefu, Pilar de la Puente, Cinzia Federico, Justin King, Danny Kohnen, Ravi Vij, Samuel Achilefu, Abdel Kareem Azab. Nanoparticle multispecific T-cell engagers for the treatment of multiple myeloma [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2018 Nov 27-30; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(4 Suppl):Abstract nr B01.

Abstract P3-10-18: Phase 1b trial of cirmtuzumab and paclitaxel for locally advanced, unresectable and metastatic breast cancer

Abstract Background: Cirmtuzumab is a humanized monoclonal antibody that targets the receptor tyrosine kinase like orphan receptor 1 (ROR1), which is expressed on poor prognosis breast cancer, ovarian cancer, other solid tumors, CLL and mantle cell lymphoma. One clinical study showed increased expression of ROR1 in breast cancers after neoadjuvant chemotherapy and a preclinical PDX breast cancer model showed cirmtuzumab and paclitaxel to have at least additive efficacy1. A recently completed Phase 1 trial of cirmtuzumab in CLL showed the antibody to be both safe and effective in inhibiting tumor cell ROR1 signaling in patients with CLL2. Methods: The primary aim of this trial was to determine the safety of cirmtuzumab and weekly paclitaxel in patients with advanced Her2 negative breast cancer based upon dose limiting toxicities (DLTs) in the first cycle of treatment. Secondary endpoints were clinical activity, pharmacokinetics and correlative biomarkers on tumor specimens. Eligible patients were those with locally advanced, unresectable or metastatic Her2 negative breast cancer who had not received paclitaxel in the metastatic setting, had not developed metastatic disease within 6 months of (neo)adjuvant paclitaxel, had ECOG performance status of 0-2, and had adequate laboratory parameters. Any number of prior lines of therapy were allowed. Study treatment included fixed dose 600 mg cirmtuzumab given days 1 and 15 of cycle 1 and then day 1 of each subsequent 28-day cycle. Paclitaxel was given weekly at a dose of 80mg/m2. Patients were evaluated in dose cohorts of 5 for DLTs with a target of 15 evaluable patients. Results: To date, 6 patients evaluable for safety and 5 patients evaluable for DLTs were treated. Age range was 30 to 59 years. Three of 6 safety-evaluable patients had triple negative breast cancer at study enrollment. Prior lines of chemotherapy in the metastatic setting ranged from 0-3. No discontinuations for toxicity and no DLTs have been observed to date. Adverse events (AEs) have been consistent with the known safety profile of paclitaxel, with 3 episodes of grade 3 neutropenia in 2 patients and 1 episode of grade 3 hyperglycemia. All other AEs were grade 1 or 2. Partial responses have been observed in 2/5 patients with one patient response ongoing with cirmtuzumab alone for at least 17 weeks after stopping paclitaxel. Pharmacokinetic analysis of serial plasma samples for free unbound antibody from two patients provided results similar to those observed in CLL patients treated with cirmtuzumabwith a projected half-life of 30 days2. No decline in antibody concentration over time was observed consistent with the absence of neutralizing antibodies. Conclusions: Preliminary information indicates that the combination of fixed dose cirmtuzumab plus paclitaxel is well-tolerated and safe. Responses to therapy have been observed and preliminary pharmacokinetic results are consistent with sustained potentially therapeutic levels. Updated safety, clinical activity, pharmacokinetics and biomarker analyses will be presented. 1Zhang S et al. Proc Natl Acad Sci USA. 116(4): 1370-1377. PMID 30622177. 2Choi MY et al. Cell Stem Cell 22(6): 951-959. PMID 29859176. Funding sources: CIRM UC San Diego Alpha Stem Cell Clinic and Sanford Stem Cell Clinical Center; Oncternal Therapeutics, Inc.; UC San Diego Moores Cancer Center Padres Pedal the Cause Grant; Gonick Breast Cancer Research Fund Citation Format: Rebecca A Shatsky, Richard B Schwab, Teresa L Helsten, Emily I Pittman, Ruifeng Chen, James B Breitmeyer, Catriona HM Jamieson, Thomas J Kipps, Barbara A Parker. Phase 1b trial of cirmtuzumab and paclitaxel for locally advanced, unresectable and metastatic breast cancer [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P3-10-18.

Maternal urinary bisphenol A concentration and thyroid hormone levels of Chinese mothers and newborns by maternal body mass index.

Animal studies indicated that bisphenol A (BPA) exposure during pregnancy may disrupt thyroid function which is critical for fetal development. However, few epidemiological studies have examined this topic and the results were inconsistent. We aimed to evaluate whether prenatal BPA exposure is associated with thyroid hormone levels in Chinese mothers and newborns with stratification by maternal body mass index (BMI). BPA concentration were measured in urine samples collected from 555 women at late pregnancy. Maternal serum free thyroxin (FT4), thyroid-stimulating hormone (TSH) and thyroid peroxidase antibody (TPO-Ab) concentrations at the third trimester were abstracted from medical records. Cord serum-free triiodothyronine (FT3), FT4, TSH, and TPO-Ab levels were measured in 398 newborns. Prenatal urinary BPA was detected in 98.5% of mothers with a geometric mean of 1.32ng/mL (95% CI 1.17-1.49ng/mL). With each 10-fold increase in BPA concentrations, maternal log10_(TSH) mIU/L was 0.10 lowered (95% CI - 0.20, - 0.005, p < 0.05) among pre-pregnancy BMI>23kg/m2, with adjustment for maternal age, maternal education, gestation diabetes mellitus (GDM), husband smoking during pregnancy, parity, and gestational age at thyroid parameters measured, but no association was observed in pre-pregnancy BMI < 18.5, or 18.5-22.9kg/m2 stratum. No BPA-associated changes were observed in maternal FT4 level or odds of positive TPO-Ab in all BMI stratum. Also, no associations were observed between prenatal urinary BPA concentration and cord serum FT4, FT3, TSH levels, and odds of positive TPO-Ab in both male and female newborns among pre-pregnancy BMI < 18.5, 18.5-22.9 or>23kg/m2 stratum. In this study, prenatal urinary BPA concentration was associated with lower maternal TSH among women with overweight, but not associated with other maternal thyroid parameters or cord serum thyroid parameters across maternal BMI categories. More research on pregnant women and newborns cohort with BPA exposure are warranted.

P216 Anti-drug antibodies to infliximab: A comparison of free anti-drug antibody measurement

Abstract Background Infliximab (IFX) is a biologic drug that inhibits the action of TNFα, a pro-inflammatory cytokine implicated in the pathogenesis of inflammatory bowel disease (IBD). IFX has greatly improved the outcomes for patients with IBD, but response to the drug is not universal. Primary non-response to IFX treatment occurs in up to 30% of IBD patients while up to 46% of patients develop secondary loss of response.1,2 Anti-drug antibodies (ADAs) to IFX are considered a significant risk factor for the loss of response to treatment, hence the measurement of ADAs as part of therapeutic drug monitoring is an increasingly utilised tool. The detection of ADAs varies widely depending on the type of assays used. The aim of this study was to determine the qualitative concordance of three commercially available ELISA kits for measurement of free ADAs to IFX on the Grifols Triturus analyser. Methods 150 patient samples with low IFX drug levels (≤0.6 µg/ml) were analysed for free ADAs using Promonitor, Lisa Tracker and IDKmonitor kits on the Grifols Triturus automated ELISA analyser. Results Kappa coefficient (κ) analysis indicated a moderate agreement between the Promonitor and IDKmonitor assays (κ = 0.484 (95% CI, 0.357 to 0.611)) and the IDKmonitor and Lisa Tracker assays (κ = 0.485 (95% CI, 0.348–0.621)) as well as substantial agreement between the Promonitor and Lisa Tracker assays (κ = 0.768 (95% CI, 0.667–0.870)). Figure 1 shows the distribution of samples identified as free ADA positive by each kit. Conclusion Although a broad qualitative agreement was found between the three kits, they should not be used interchangeably for patient management. All kits appear amenable for utilisation in a high-throughput laboratory though a true quantitative comparison between these kits was precluded by the absence of any certified reference material for free ADAs to IFX. Further research is required to estimate the impact of free ADAs on efficiency of IFX treatment and patient management. References

Features of humoral immunity in cows infected with the leukaemia virus

The article presents the results of studying the dynamics of the formation of antibodies and immune complexes, reveals the prospects for improving the early diagnosis of cattle leukaemia. Studies were conducted for 6 months on an experimental group of animals consisting of 12 cows. The titers of free and bound antibodies in blood serum and milk were determined by enzyme-linked immunosorbent assay. The results of studies showing that changes in titers of anti-leukaemia antibodies in the blood serum of cows naturally infected with BLV (bovine leukaemia virus) are significantly different from experimental infection data are adduced. In cows infected with BLV, there is no definite relationship between antibody titers in milk and in blood serum. With sufficiently high titers of serum antibodies, antibody titers in milk can be minimal for the same cows; conversely, with low titers of serum antibodies, there can be high antibody titers in milk. In the titers of antibodies free and bound in the immune complexes in blood serum with the development of the disease, a certain dependence is traced. With a decrease in titers of free antibodies, in most cases, an increase in titers of “bound” antibodies is observed, i.e., the degree of formation of circulating immune complexes (CECs) increases. There is no clearly defined dependence in the dynamics of changes in titers free and bound in immune complexes of antibodies in milk samples. They can remain at the same level for a long time, both at low and rather high levels.

Evaluation of Reproductive Outcome in Infertile Hypothyroid Women on Thyroxine Therapy.

Introduction:Thyroid dysfunction is associated with increased risk of infertility. Serum thyroid stimulating hormone (TSH) screening in all women seeking infertility care is recommended and hypothyroid infertile women should be treated with thyroxine until the preconception serum TSH level is <2.5 mU/l.[1] However, insufficient evidence exist to determine if thyroxine therapy improves fertility in subclinical hypothyroid women who are trying to conceive naturally.Objectives:The objective is to study the effect of thyroxine therapy on reproductive outcome in infertile women with clinical and subclinical hypothyroidism (SCH).Materials and Methods:The study is a descriptive cohort study with 72 subjects. Women between 20 and 40 years of age with primary or secondary infertility with hypothyroidism were studied and thyroid profile including free T3, T4, TSH, and thyroid antibodies were done. Thyroxine was given to clinical, subclinical hypothyroid subjects depending on TSH levels such that serum TSH levels are maintained < 2.5 mU/L. Serial thyroid function test was done every 6 weeks until the optimal levels were reached. Once normal TSH levels were reached subjects were followed up for 6 months. Reproductive outcome was analyzed in two groups. Group A included hypothyroid infertile women who conceived and Group B included those who did not conceive following thyroxine therapy.Results:Thirty-eight out of 72 subjects (54%) conceived during thyroxine treatment (Group A) of which 4 cases had miscarriage. Maximum infertile women in Group A (20/38) conceived between 6 and 12 months (52.6%) of thyroxine therapy. Significant decrease was observed in mean TSH levels over a period of 6 months (P < 0.001). The infertility period until pregnancy in Group A reduced significantly from 5.2 ± 1.8 years to 0.5 ± 0.8 years after thyroxine treatment (P = 0.001).Conclusion:Thyroxine therapy enhances fertility in infertile women with clinical and SCH.

Highly sensitive lateral flow test with indirect labelling for zearalenone in baby food

ABSTRACT Maximum permissible levels of mycotoxins in baby food may be 1% of those in ordinary food. Therefore, highly sensitive methods of mycotoxin control are in demand. To detect such low amounts, expensive instrumental methods are commonly used. Advantages of immunochromatographic analyses are their low cost and simple sample preparation; however, their sensitivity needs to be increased to contend with instrumental methods. A scheme for competitive immunochromatography with indirect labelling was implemented and developed for the detection of mycotoxin zearalenone (ZEA). Two separate reagents were used for the assay, namely free specific antibodies and antispecies antibodies conjugated with gold nanoparticles. This made it possible to simultaneously increase the sensitivity of the assay and the reliability of measurements. The instrumental detection limit of ZEA in baby food was 5 pg/mL (100 pg/g). Thus, the sensitivity attained is comparable with liquid chromatography characteristics. The duration of the analysis was 17 min.

Steroid-sparing effect of omalizumab in stage IV (corticosteroid dependent) allergic bronchopulmonary aspergillosis

Allergic bronchopulmonary aspergillosis (ABPA) consists of five stages ranging from the acute stage (pulmonary infiltrate, eosinophilia, and increased total Immunoglobulin-E [IgE]) to pulmonary fibrosis. Our patients were diagnosed as Stage IV (corticosteroid dependent) ABPA, in which treatment with systemic corticosteroids cannot be discontinued as symptoms worsen after discontinuation of oral corticosteroids. Omalizumab is a humanized recombinant, monoclonal antibody that selectively binds to IgE to inhibit the immune response to an allergen. It forms immune complexes which not only lower the level of free IgE antibody, but also serve as an allergen buffer, even in cases of markedly high serum IgE >1000 IU/ml in ABPA. In this article, we report three cases of (Stage IV) corticosteroid-dependent ABPA with poor clinical response even after effective treatment with antifungal therapy and courses of corticosteroids (>3 exacerbations/year). After the start of injection omalizumab, the three patients have experienced significant and sustained clinical improvement and improved quality of life with low-dose prednisolone 5 mg every alternate day. The dose of systemic corticosteroids was reduced by 50% in the 1st year and by 70% in the 2nd year although lung function failed to improve.

Effect of oral isotretinoin therapy on thyroid function in patients with moderate-to-severe acne vulgaris: A prospective study

Introduction: Despite its growing use, the side effects of isotretinoin therapy are not fully understood. Studies have suggested that isotretinoin can cause thyroid dysfunction, suggesting the need to investigate thyroid dysfunction as a possible side effect of oral isotretinoin therapy. Purpose: The main objective of the study is to investigate the association between isotretinoin therapy and thyroid dysfunction. Methods: In all, 51 patients (male, 21; female, 30; aged 18–25 years) with moderate-to-severe acne vulgaris were treated with 0.5 mg/kg/day oral isotretinoin (cumulative dose: 120–150 mg/kg). Serum-free thyroxine (T4), thyroid-stimulating hormone (TSH), and thyroglobulin (TGA) and thyroid peroxidase (TPO) antibody levels were measured. Results: TSH levels at baseline and at the 2nd, 4th, 6th, and 8th months of treatment were 2.3307 ± 1.097, 2.5824 ± 1.34, 2.678 ± 1.133, 2.3 ± 1.17, and 2.7 ± 2.17 IU·mL, respectively, with no significant change from baseline (P = 0.526, P = 0.552, P = 0.530, and P = 0.910 at 2, 4, 6, and 8 months, respectively). Serum levels of free T4, TGA, and TPO antibodies were unchanged. Conclusions: Follow-up screening for thyroid dysfunction may not be necessary for patients receiving oral isotretinoin.

Oral delivery of anti-TNF antibody shielded by natural polyphenol-mediated supramolecular assembly for inflammatory bowel disease therapy.

Rationale: Anti-tumor necrosis factor (TNF) therapy is a very effective way to treat inflammatory bowel disease. However, systemic exposure to anti-TNF-α antibodies through current clinical systemic administration can cause serious adverse effects in many patients. Here, we report a facile prepared self-assembled supramolecular nanoparticle based on natural polyphenol tannic acid and poly(ethylene glycol) containing polymer for oral antibody delivery.Method: This supramolecular nanoparticle was fabricated within minutes in aqueous solution and easily scaled up to gram level due to their pH-dependent reversible assembly. DSS-induced colitis model was prepared to evaluate the ability of inflammatory colon targeting ability and therapeutic efficacy of this antibody-loaded nanoparticles.Results: This polyphenol-based nanoparticle can be aqueous assembly without organic solvent and thus scaled up easily. The oral administration of antibody loaded nanoparticle achieved high accumulation in the inflamed colon and low systemic exposure. The novel formulation of anti-TNF-α antibodies administrated orally achieved high efficacy in the treatment of colitis mice compared with free antibodies administered orally. The average weight, colon length, and inflammatory factors in colon and serum of colitis mice after the treatment of novel formulation of anti-TNF-α antibodies even reached the similar level to healthy controls.Conclusion: This polyphenol-based supramolecular nanoparticle is a promising platform for oral delivery of antibodies for the treatment of inflammatory bowel diseases, which may have promising clinical translation prospects.

Mathematical analysis of an HBV model with antibody and spatial heterogeneity.

In this paper, we modify the HBV model proposed in [1] to include the spatial variations of free antibody, virus-antibody complexes, and free virus. By using comparison arguments and theory of uniform persistence, we can show that the persistene/extinction of HBV can be determined by the reproduction number(s).

Detection by Flow Cytometry of Anti-DNA Autoantibodies and Circulating DNA Immune Complexes in Lupus Erythematosus

A new method for the detection by flow cytometry of anti-double-stranded DNA antibodies and of circulating immune complexes (IC) containing endogenous DNA (IC-eDNA) is described. From each serum sample, two samples were taken, one was used to detect IC-eDNA. The other to detect anti-DNA antibodies was incubated with calf thymus DNA. ICs were isolated by polyethylene glycol precipitation or by cryoprecipitation, after which immunoglobulins were labeled with FITC-conjugated anti-human globulin. Serum samples from 63 systemic lupus erythematosus (SLE) patients, 32 incomplete lupus, and 87 control patients were tested. Detection of anti-dsDNA antibodies by flow cytometry had a diagnostic sensitivity and specificity almost comparable to routine tests, the fluorescent enzyme immunoassay EliA™-dsDNA test, and the ultrasensitive Crithidia luciliae indirect immunofluorescence test. In 21 (33%) out of 63 SLE serum samples, IC-eDNA was detected. In these samples, free anti-dsDNA antibodies were hardly detectable or undetectable by flow cytometry or by routine tests. When anti-DNA antibodies are neutralized by endogenous DNA and can no longer be detected by routine tests, the serologic diagnosis and the follow-up of relapses in patients with SLE is compromised. To overcome this obstacle, we propose an accessible solution: the detection of circulating IC-eDNA by flow cytometry.

Hook effect detection and detection-range-controllable one-step immunosensor for inflammation monitoring

Immune reactions are affected by the hook effect, which can cause false-negative results at high concentrations. We introduce a hook effect detecting and controllable real one-step immunoassay that can be used to measure a target analyte quantitatively in undiluted clinical samples. The target analyte examined is canine C-reactive protein (cCRP), which is a major acute phase protein in dogs. The serum concentration of this protein changes in response to systemic inflammation. cCRP is an immune analyte that is measured by an antigen–antibody reaction. The cCRP assay is more vulnerable to the hook effect owing to extremely high concentrations compared to other immune analytes. We describe a process that can detect the hook effect using free antigen and can control the detection range using free antibody on cCRP antibody-labeled gold nanoparticles with high performance. The hook effect was not observed at extremely high concentrations of cCRP, and the detection range was dramatically increased.