Role of alpha‐1 antitrypsin in human health and disease

Alpha‐1 antitrypsin (AAT) deficiency is an under‐recognized hereditary disorder associated with the premature onset of chronic obstructive pulmonary disease, liver cirrhosis in children and adults, and less frequently, relapsing panniculitis, systemic vasculitis and other inflammatory, autoimmune and neoplastic diseases. Severe AAT deficiency mainly affects Caucasian individuals and has its highest prevalence (1 : 2000–1 : 5000 individuals) in Northern, Western and Central Europe. In the USA and Canada, the prevalence is 1: 5000–10 000. Prevalence is five times lower in Latin American countries and is rare or nonexistent in African and Asian individuals. The key to successful diagnosis is by measuring serum AAT, followed by the determination of the phenotype or genotype if low concentrations are found. Case detection allows implementation of genetic counselling and, in selected cases, the application of augmentation therapy. Over the past decade, it has been demonstrated that AAT is a broad‐spectrum anti‐inflammatory, immunomodulatory, anti‐infective and tissue‐repair molecule. These new capacities are promoting an increasing number of clinical studies, new pharmacological formulations, new patent applications and the search for alternative sources of AAT (including transgenic and recombinant AAT) to meet the expected demand for treating a large number of diseases, inside and outside the context of AAT deficiency.     

Discrimination between COPD patients with and without alpha 1-antitrypsin deficiency using an electronic nose

Background and objective: To compare the volatile organic compound patterns of patients with COPD with and without alpha 1‐antitrypsin (AAT) deficiency using electronic nose technology. Methods: Exhaled breath condensate and pure exhaled breath of patients with COPD with (n = 10) and without (n = 23) AAT deficiency and healthy controls (n = 10) were analysed. The effect of human recombinant AAT on the volatile organic compound profile of 11 AAT‐deficient patients was also examined. Exhaled breath condensate and pure exhaled breath were measured using the Cyranose 320. Smell prints were analysed by linear discriminant analysis (LDA) using Mahalanobis distance (MD) and cross‐validation values (CVVs). Results: Smell prints of patients with AAT‐deficiency were different from those with COPD in exhaled breath condensate (LDA: P < 0.0001, sensitivity of 1.00, specificity of 1.00, CVV 82.0%, MD 2.37) and in pure exhaled breath (LDA: P < 0.0001, sensitivity of 1.00, specificity of 1.00, CVV 58.3%, MD 2.27). Smell prints of AAT‐deficient patients before and after human recombinant AAT augmentation were different (LDA: P = 0.001, sensitivity of 1.00, specificity of 1.00, CVV 53.3%, MD 1.79). Conclusions: An electronic nose can detect differences in smell prints of COPD patients with and without AAT deficiency. Augmentation therapy changes the volatile organic compound pattern. The electronic nose may be helpful in the diagnosis of AAT deficiency.     

Safety and efficacy of recombinant alpha(1)-antitrypsin therapy in cystic fibrosis

Neutrophil elastase (NE) is thought to be the most important protease which damages the cystic fibrosis (CF) lung. Attempts have been made to suppress this activity using the plasma-derived inhibitor, alpha(1)-antitrypsin (AAT). In this pilot study, the safety and efficacy of inhaled recombinant human AAT (rAAT) as a treatment for CF were investigated. Thirty-nine patients participated in a prospective, double-blinded, randomized, placebo-controlled phase II trial to examine the effect of rAAT (500, 250, and 125 mg) on sputum NE activity. Sputum myeloperoxidase (MPO), interleukin-8, tumor necrosis factor receptors, sputum and plasma NE/AAT complexes, and safety parameters were also measured. Subjects were randomized to receive nebulized treatment once a day for 4 weeks, followed by 2-4 weeks with no study treatment, and then a 2-week rechallenge phase. Trends toward a reduction in NE activity were observed in patients treated with 500 mg and 250 mg of rAAT compared to placebo. Sputum NE/AAT complex and MPO levels were lower on rAAT compared to placebo. No major adverse events and, in particular, no allergic reactions to rAAT were observed. Although significant differences between rAAT and placebo for sputum NE activity were not observed, some improvements were found for secondary efficacy variables. This study demonstrated that nebulized rAAT is safe and well-tolerated, but has a limited effect on NE activity and other markers of inflammation.     

Validation and development of an immunonephelometric assay for the determination of alpha-1 antitrypsin levels in dried blood spots from patients with COPD

OBJECTIVE:

To validate and develop an immunonephelometric assay for the determination of alpha-1 antitrypsin (AAT) levels in dried blood spots from COPD patients in Brazil.

METHODS:

We determined AAT levels in serum samples and dried blood spots from 192 COPD patients. For the preparation of dried blood spots, a disk (diameter, 6 mm) was placed into a tube, eluted with 200 µL of PBS, and stored overnight at 4ºC. All of the samples were analyzed by immunonephelometry in duplicate. We used the bootstrap resampling method in order to determine a cut-off point for AAT levels in dried blood spots.

RESULTS:

The correlation coefficient between the AAT levels in serum samples and those in dried blood spots was r = 0.45. For dried blood spots, the cut-off value was 2.02 mg/dL (97% CI: 1.45-2.64 mg/dL), with a sensitivity, specificity, positive predictive value, and negative predictive value of 100%, 95.7%, 27.2%, and 100%, respectively.

CONCLUSIONS:

This method for the determination of AAT levels in dried blood spots appears to be a reliable screening tool for patients with AAT deficiency.     

Calibration of the second International Standard for hepatitis B immunoglobulin in an international collaborative study

Background and Objectives The International Standard for hepatitis B immunoglobulin is used in the standardization of the anti‐HBs content of immunoglobulins for prophylactic and therapeutic use and also in the standardization and calibration of quantitative diagnostic anti‐HBs assay kits. A collaborative study was undertaken to assess the suitability of a candidate Second International Standard (2nd IS), and to calibrate it in International Units (IU). Materials and Methods The candidate 2nd IS was prepared from a bulk of 5% hepatitis B immunoglobulin (NIBSC code 07/164). Twenty‐two participants from 12 countries assayed the first IS, the candidate 2nd IS, a freeze‐dried pool of plasma containing anti‐HBs and a plasma from a blood donor. These samples were assayed with 19 different assay kits. Results Data from 102 assays were received. The mean potencies of two coded samples of the candidate 2nd IS were 100·7 and 101·4 IU/ml (combined potency 101·0 IU/ml). The geometric coefficients of variation for these samples were both 13%. The predicted long‐term stability of 07/164 was assessed by assaying samples stored at elevated temperatures for a period of 6 months. 07/164 was predicted to be stable at −20°C with the estimated % loss per year of below 0·2%. Conclusion 07/164 was established as the 2nd IS for hepatitis B immunoglobulin with an assigned potency of 100 IU/ampoule by the WHO Expert Committee on Biological Standardisation. The United States Food and Drug Administration has adopted the same standard as the new Reference for Hepatitis B Immunoglobulin, Lot 3.     

An association between Type 2 diabetes and α1‐antitrypsin deficiency

Aims α₁‐Antitrypsin (AAT) is a serine protease inhibitor which recently has been shown to prevent Type 1 diabetes development, to prolong islet allograft survival and to inhibit pancreatic B‐cell apoptosis in vivo. It has also been reported that Type 1 diabetic patients have significantly lower plasma concentrations of AAT, suggesting the potential role of AAT in the pathogenesis of Type 1 diabetes. We have investigated whether plasma AAT levels are altered in Type 2 diabetes. Methods The study included patients with Type 2 diabetes (n = 163) and non‐diabetic control subjects matched for age, sex and smoking habits (n = 158) derived from the population‐based Malmö Diet and Cancer study. Plasma samples were analysed for AAT concentration and phenotype and serum glucose, insulin, C‐reactive protein and lipid levels were measured. Glycated haemoglobin was also measured. Results In the diabetic group, the women had higher mean plasma AAT levels than men (P < 0.05). The mean plasma AAT levels did not differ between diabetic and control subjects. However, the number of individuals with low AAT levels (< 1.0 mg/ml) was 50% higher in the diabetic group (P < 0.05) and the frequency of AAT deficiency genotypes was 50% higher (NS) in diabetic compared with control subjects. In the group of diabetic patients with AAT < 1 mg/ml, AAT directly correlated with systolic blood pressure (P = 0.048) and inversely correlated with waist–hip ratio (P = 0.031). Conclusions Our results provide evidence that deficiency of AAT may be associated with an increased risk of developing Type 2 diabetes.     

The First International Standard for Insulin-like Growth Factor-1 (IGF-1) for immunoassay: Preparation and calibration in an international collaborative study

The World Health Organization (WHO) Expert Committee on Biological Standardization (ECBS) has recognized the need for an International Standard for Insulin-like Growth Factor-1 (IGF-1) for the calibration of immunoassays and for the monitoring of the content of therapeutic products. The objective of the study reported here was the characterization of a candidate standard for IGF-1 in an international collaborative study carried out by 18 laboratories in nine countries, by comparison with (i) a primary calibrant characterized by amino acid analysis and UV spectroscopy, and (ii) the existing International Reference Reagent coded 87/518 by HPLC, immunoassay and bioassay. The study was designed as follows: Phase I involved the establishment of a primary calibrant of rhIGF-1, containing approximately 1.0 mg rhIGF-1 per vial. A defined value was assigned to the primary calibrant by amino acid analysis (AAA) and UV spectroscopy. Phase II involved calibration of the candidate standard in terms of the primary calibrant by HPLC, with confirmatory data from immunoassay and bioassay. Results from Phase I confirmed the primary calibrant as containing 1.045 mg per vial. Although there was some variability among laboratory estimates of IGF-1 in the proposed standard using the different methods in Phase II, the estimates by the various methods were in broad agreement. On the basis of the results reported here, the World Health Organization (WHO) has established the preparation coded 02/254 as the First International Standard for Insulin-like Growth Factor-1, human, recombinant, for immunoassay with an assigned content of 8.50 μg per ampoule. Details of how to order the standard can be found at www.nibsc.ac.uk.     

Exhaled nitric oxide levels in alpha‐1‐antitrypsin PiMZ subjects

Objectives. Although the likelihood of intermediate alpha‐1‐antitrypsin deficiency (PiMZ) patients developing chronic obstructive pulmonary disease (COPD) remains uncertain, several investigators have suggested that a lack of antiprotease inhibitor activity may favour the development of airway inflammation with subsequent pulmonary tissue damage. The levels of exhaled nitric oxide (FeNO) in PiMZ subjects are unknown and polymorphisms in nitric oxide synthase have been linked to lung disease susceptibility in subjects with alpha‐1‐antitrypsin (AAT) deficiency. This study was aimed at assessing FeNO levels in a group of PiMZ subjects and comparing it with the concentrations found amongst groups of COPD and control patients. Design. A group of 31 PiMZ subjects, 31 COPD patients and 30 controls underwent pulmonary function tests, AAT assay and phenotyping, and FeNO measurement in an ambulatory setting. Results. FeNO values observed in the group of PiMZ subjects (21.6 ± 8.9 ppb) showed a significant increase compared with COPD (14.5 ± 8.7 ppb; P < 0.01) and the control groups (9.1 ± 2.9 ppb; P < 0.01). Within the PiMZ population, a significant, negative correlation was observed between plasma AAT levels and FeNO readings. Conclusions. Not only did PiMZ subjects show increased FeNO levels compared with COPD patients and controls; FeNO levels proved to be related to the reduced concentration of plasma AAT. Such findings seem to suggest the importance of FeNO measurements on PiMZ subjects for monitoring a possible progression of airway inflammation to obstructive lung disease as observed in some of these patients.     

An international collaborative study to replace the 1st international standard for prekallikrein activator

BACKGROUND AND OBJECTIVES: The aim of this study was to replace the 1st International Standard (IS) for prekallikrein activator (PKA) (code 82/530) with a new IS and European Biological Reference Preparation (BRP). The new standards were freeze dried 20% albumin solution containing PKA, the same solution that is tested using these standards. Aspects of the methodology for PKA determination were also examined as part of this study. MATERIALS AND METHODS: A batch of 20% albumin containing approximately 30 IU/ml was donated by a manufacturer of blood products and dispensed into ampoules at the National Institute for Biological Standards and Control (NIBSC) to create the candidate IS (02/168, sample B in the study) and at the European Directorate for the Quality of Medicines (EDQM) to create the candidate BRP (sample C in the study). The concentration of PKA in these preparations was determined in an international collaborative study involving 31 laboratories from 17 countries worldwide in comparison with the 1st IS for PKA (82/530) containing 85 IU of PKA per ampoule. Participants were requested to perform their own in-house method, based on the current Ph. Eur. monograph for determination of PKA in albumin solutions. Participants were provided with sufficient samples to perform two or three assays and were asked to use their local prekallikrein substrate (PKS) and also to use a commercial PKS provided as part of the study, in order to investigate the importance of the source of PKS on the final potency values. RESULTS: Samples B and C emerged with identical PKA concentrations of 29 IU/ml, very close to the expected value. This figure was determined using a variety of statistical methods, with the participants' own calculated values and values calculated centrally at the EDQM using raw data. The value of 29 IU/ml was consistent and independent of the method of calculation, although interlaboratory variability was more sensitive to the statistical analysis method. There was no statistically significant difference in mean potencies when comparing results with the laboratories' own local substrate and the substrate provided for the study. All stability studies indicate that these lyophilized preparations of PKA in 20% albumin are extremely stable. CONCLUSIONS: Samples B and C were established as the 2nd IS (code 02/168) and PKA activator in albumin BRP batch 1 (Y0000263), respectively, with a potency of 29 IU per ampoule. Results from this study indicate that testing for PKA in albumin may be less sensitive to the source of PKS than previously feared. The study highlights a number of methodological issues that may need revising in the Ph. Eur. general chapter 2.6.15.     

Diagnosis and treatment of lung disease associated with alpha one‐antitrypsin deficiency: A position statement from the Thoracic Society of Australia and New Zealand*

AATD is a common inherited disorder associated with an increased risk of developing pulmonary emphysema and liver disease. Many people with AATD‐associated pulmonary emphysema remain undiagnosed and therefore without access to care and counselling specific to the disease. AAT augmentation therapy is available and consists of i.v. infusions of exogenous AAT protein harvested from pooled blood products. Its clinical efficacy has been the subject of some debate and the use of AAT augmentation therapy was recently permitted by regulators in Australia and New Zealand, although treatment is not presently subsidized by the government in either country. The purpose of this position statement is to review the evidence for diagnosis and treatment of AATD‐related lung disease with reference to the Australian and New Zealand population. The clinical efficacy and adverse events of AAT augmentation therapy were evaluated by a systematic review, and the GRADE process was employed to move from evidence to recommendation. Other sections address the wide range of issues to be considered in the care of the individual with AATD‐related lung disease: when and how to test for AATD, changing diagnostic techniques, monitoring of progression, disease in heterozygous AATD and pharmacological and non‐pharmacological therapy including surgical options for severe disease. Consideration is also given to broader issues in AATD that respiratory healthcare staff may encounter: genetic counselling, patient support groups, monitoring for liver disease and the need to establish national registries for people with AATD in Australia and New Zealand.     

Actualización sobre indicaciones de búsqueda activa de casos y tratamiento con alfa-1 antitripsina por vía intravenosa en pacientes con enfermedad pulmonar obstructiva crónica asociada a déficit de alfa-1 antitripsina

The effect of hereditary alpha-1 antitrypsin (AAT) deficiency can manifest clinically in the form of chronic obstructive pulmonary disease (COPD). AAT deficiency (AATD) is defined as a serum concentration lower than 35% of the expected mean value or 50 mg/dl (determined by nephelometry). It is associated in over 95% of cases with Pi*ZZ genotypes, and much less frequently with other genotypes resulting from combinations of Z, S, rare and null alleles. A systematic qualitative review was made of 107 articles, focusing mainly on an active search for AATD in COPD patients and intravenous (iv) treatment with AAT. On the basis of this review, the consultant committee of the Spanish Registry of Patients with AATD recommends that all COPD patients be screened for AATD with the determination of AAT serum concentrations, and when these are low, the evaluation must be completed with phenotyping and, on occasions, genotyping. Patients with severe AATD COPD should receive the pharmacological and non-pharmacological treatment recommended in the COPD guidelines. There is enough evidence from large observational studies and randomized placebo-controlled clinical trials to show that the administration of iv AAT reduces mortality and slows the progression of emphysema, hence its indication in selected cases that meet the inclusion criteria stipulated in international guidelines.The administration of periodic infusions of AAT is the only specific treatment for delaying the progression of emphysema associated with AATD.     

Recombinant factor IX: discrepancies between one‐stage clotting and chromogenic assays

New and modified recombinant factor IX (rFIX) products are in development and accurate potency estimation is important to ensure the consistency of production and efficacy of these therapeutics. Collaborative study data obtained during the replacement of the 3rd International Standard (IS) for FIX concentrate suggested that there was a discrepancy between potency estimates for rFIX using clotting and chromogenic methods, when the rFIX candidate was measured against the plasma‐derived FIX (pdFIX) IS. This study explores potential chromogenic and one‐stage clotting method discrepancies in more detail. Five batches each of rFIX and pdFIX were assayed against the 4th IS FIX concentrate (a pdFIX) by activated partial thromboplastin time (APTT) one‐stage clotting assay and specific functional chromogenic assay. The potency of rFIX by chromogenic assay was consistently around 70% of the one‐stage clotting potency (average 78 and 108 IU mL^?1 respectively). These differences were not observed with pdFIX, which had similar potencies (average 96 IU mL^?1) by each assay method. In addition, different APTT reagents yielded different potency estimates for rFIX when assayed against the pdFIX IS, with a variation of up to 23%. In all cases, the differences were largely resolved when a rFIX reference was used as the standard. This study highlights some of the challenges associated with assay of rFIX products in the laboratory and that careful consideration needs to be given to the choice of reference material used. This is especially important with the imminent arrival of new and modified rFIX products.     

von Willebrand factor standards for plasma and concentrate testing

The wide range of plasma von Willebrand factor (vWF) levels in the normal population makes vWF an ideal candidate for calibration in World Health Organization International Standards (WHO IS). Calibration of the WHO IS Plasma for vWF:antigen (vWF:Ag), vWF:ristocetin cofactor activity (vWF:RCo), and vWF:collagen-binding activity (vWF:CB) addresses the need to reconcile the original definition of the International Unit (IU) in fresh normal plasma with continuity between replacement freeze-dried preparations, whereas calibration of the WHO IS Concentrate aims to maintain equivalence between the IU used for the estimation of vWF in plasma (diagnosis) and the IU used for the potency labeling of therapeutic concentrates (replacement therapy). The latter objective was achieved through the calibration of the WHO 1st IS vWF Concentrate, for vWF:Ag and vWF:RCo, by assay directly relative to the WHO IS Plasma. Calibration of the WHO IS Concentrate for vWF:CB was not possible due to large interlaboratory and intermethod discrepancies, and resolution of this problem may require the use of recommended methodology. There is evidence that the use of the WHO IS Concentrate leads to considerable reduction in interlaboratory variability for vWF estimates in therapeutic concentrates and hence the "assay like versus like" principle should be applied.     

An international collaborative study to establish a replacement World Health Organization International Standard for human immunodeficiency virus 1 RNA nucleic acid assays

Background and Objectives An international collaborative study was undertaken to identify a replacement for the World Health Organization (WHO) 1st International Standard for human immunodeficiency virus 1 (HIV‐1) RNA for use in nucleic acid‐based techniques (NAT) (code 97/656). In the original study to establish the 1st International Standard, a second candidate material (code 97/650) had been shown to perform well and this was re‐evaluated to establish whether it would be a suitable replacement. Materials and Methods Eight laboratories from six different countries participated in the collaborative study to evaluate the candidate replacement standard. A total of eight different NATs were used, five in a quantitative format and three qualitative, of which five were commercially available. Results The results showed that the estimates of RNA copies in the current study were generally in line with those of the original study and there was no evidence of any drift in overall levels expressed in International Units (IU) for the candidate standard between the two studies. Furthermore, it was shown to be stable over long‐term storage at –20°C. Conclusions The candidate material code 97/650 was established by the WHO as the 2nd International Standard for HIV‐1 RNA for use in NAT and assigned a unitage of 5·56 log₁₀ (363 078) IU/vial.     

曼氏裂头蚴cDNA文库的构建及诊断候选抗原筛选

目的　构建曼氏裂头蚴cDNA文库,通过免疫筛选获得曼氏裂头蚴病诊断候选抗原基因。方法　提取曼氏裂头蚴虫体总RNA,反转录合成cDNA,连接噬菌体载体,经体外包装后构建曼氏裂头蚴SMART cDNA文库。用曼氏裂头蚴病患者血清免疫筛选cDNA文库,获得阳性克隆,测定阳性克隆插入片段的DNA序列,进行同源性分析并预测编码蛋白的结构和功能。结果　成功构建了曼氏裂头蚴cDNA文库,文库滴度为6.25 × 106 pfu/mL,重组率为100%,文库插入片段的平均长度大于1 100 bp。经免疫筛选获得12个阳性克隆,分为Sm⁃Ⅰ、Sm⁃Ⅱ、Sm⁃Ⅲ和Sm⁃Ⅳ4类,其代表克隆为Sm60⁃1、Sm58⁃1、Sm20⁃1、Sm22⁃3,插入片段长度分别为1 134、1 063、883、969 bp,编码蛋白分别与欧猥迭宫绦虫抗原多肽、胞质抗原、核糖体蛋白S4样蛋白和未命名蛋白具有较高同源性。结论　成功构建了曼氏裂头蚴SMART cDNA文库,获得了4类阳性克隆,为曼氏裂头蚴病诊断抗原的进一步研究奠定了基础。     

High Correlation Between Clearance of Renal Protein‐Bound Uremic Toxins (Indoxyl Sulfate and p‐Cresyl Sulfate) and Renal Water‐Soluble Toxins in Peritoneal Dialysis Patients

Peritoneal dialysis (PD) is characterized by a slow continuous removal of solutes. Traditionally, dialysis adequacy is quantified by referring to the kinetics of urea nitrogen (UN) and creatinine (Cr) clearance. The efficacy of middle molecular substances and protein‐bound solutes as markers for peritoneal dialysis adequacy is not clear. The aim of this cross‐sectional study was to investigate correlations between the clearance of indoxyl sulfate (IS), p‐cresyl sulfate (PCS), UN, and Cr in the peritoneum and kidneys and to compare the overall clearances of IS and PCS between non‐anuric and anuric groups in PD patients. We recruited a total of 175 patients who had been undergoing continuous ambulatory PD (CAPD) or automated PD (APD) for at least 4 months. We measured total IS and PCS concentrations in serum, dialysate, and urine samples. Free IS and PCS concentrations were measured in all serum samples. IS and PCS clearances via both kidney and peritoneum were measured. The mean concentration of IS in the urine samples was 9.2‐fold higher than that in the dialysate samples, and concentration of PCS in the urine samples was 8.5‐fold higher than that in the dialysate samples. Peritoneal UN and Cr clearances were not correlated with peritoneal PCS clearance (P > 0.05) but were mildly correlated with peritoneal IS clearance. The peritoneal IS and PCS clearances in the different peritoneal equilibration test groups were similar. The renal UN and Cr clearances were strongly correlated with renal PCS and IS clearances (P > 0.89, P < 0.001). In addition, non‐anuric patients showed better elimination of total PCS (10.3 mg/day [range, 1.6–19.8] vs. 5.2 mg/day [range, 0–14]; P < 0.001] and IS (37.9 mg/day [range, 25.6–56.7] vs. 24.8 mg/day [range, 17.1–41.6]; P < 0.001) than anuric patients. This cross‐sectional study showed that peritoneal clearance of water‐soluble solutes is not correlated with that of PCS but is mildly correlated with that of IS. However, the renal clearances of IS and PCS show strong positive correlation with the renal clearances of UN and Cr. This study confirms the important role of residual renal function in the removal of protein‐bound uremic toxins.     

Endobronchial valve deployment in severe α‐1 antitrypsin deficiency emphysema: a case series

Introduction: Patients with end‐stage emphysema because of α‐1 antitrypsin (AAT) deficiency represent a challenging clinical management problem, and studies of volume reduction therapy to date have largely excluded these patients. We report the outcome of bronchoscopic volume reduction with the insertion of Emphasys endobronchial valves (Emphasys Medical, Redwood City, CA, USA) in six patients with end‐stage emphysema because of AAT deficiency. Case Series: Of 51 patients with end stage emphysema referred for transplantation, we studied six patients with AAT deficiency and utilized the BODE index and lung allocation score for survival estimation. Measurements and Main Results: The forced expiratory volume in 1 s improved from a median of 0.575 L to 0.905 L (P = 0.028). There was a median reduction in total lung capacity (TLC) of 0.61 L. The residual volume /TLC fell from 74.0% to 58.4%. Before treatment, four patients had a BODE index of greater than eight units, which correlates with a 4‐year survival of 18%. After treatment, two patients improved their BODE index to below seven units, which correlates with an estimated 4‐year survival of over 50%. Conclusions: The data from this case series suggest that this intervention may provide bridging therapy to subsequent transplantation for younger AAT patients with end‐stage emphysema. Please cite this paper as: Tuohy MM, Remund KF, Hilfiker R, Murphy DT, Murray JG and Egan JJ. Endobronchial valve deployment in severe α‐1 antitrypsin deficiency emphysema: a case series. Clin Respir J 2013; 7: 45–52.     

Complementation of recombinant baculoviruses by coinfection with wild-type virus facilitates production in insect larvae of antigenic proteins of hepatitis B virus and influenza virus.

We describe the coinfection of insects with wild-type and recombinant baculoviruses in which the polyhedrin gene promoter is used to express hepatitis B virus envelope protein (hepatitis B virus surface antigen; HBsAg) or influenza A virus neuraminidase (NA). Viruses were administered per os to larvae of the cabbage looper, Trichoplusia ni, causing an infection that within 5 days resulted in the production of approximately 0.15 mg of HBsAg per insect, representing 1.5% of the total extracted protein, or approximately 2.8 mg of NA per insect, representing 28% of the total extractable protein. The HBsAg and NA produced by infected larvae were purified from insect lysates. These proteins were antigenic as determined by conformation-dependent immunoassays. The NA was enzymatically active with conventional substrates. The method of infection described allows genetic complementation by wild-type virus of recombinant viruses lacking the polyhedrin gene essential for infection per os and has implications for the high-yield production in insect larvae of other recombinant proteins of baculoviruses.