Host cell protein testing by ELISAs and the use of orthogonal methods.

Host cell proteins (HCPs) are among the process-related impurities monitored during recombinant protein pharmaceutical process development. The challenges of HCP detection include (1) low levels of residual HCPs present in large excess of product protein, (2) the assay must measure a large number of different protein analytes, and (3) the population of HCP species may change during process development. Suitable methods for measuring process-related impurities are needed to support process development, process validation, and control system testing. A multi-analyte enzyme-linked immunosorbent assay (ELISA) is the workhorse method for HCP testing due to its high throughput, sensitivity and selectivity. However, as the anti-HCP antibodies, the critical reagents for HCP ELISA, do not comprehensively recognize all the HCP species, it is especially important to ensure that weak and non-immunoreactive HCPs are not overlooked by the ELISA. In some cases limited amount of antibodies to HCP species or antigen excess causes dilution-dependent non-linearity with multi-product HCP ELISA. In our experience, correct interpretation of assay data can lead to isolation and identification of co-purifying HCP with the product in some cases. Moreover, even if the antibodies for a particular HCP are present in the reagent, the corresponding HCP may not be readily detected in the ELISA due to antibody/antigen binding conditions and availability of HCP epitopes. This report reviews the use of the HCP ELISA, discusses its limitations, and demonstrates the importance of orthogonal methods, including mass spectrometry, to complement the platform HCP ELISA for support of process development. In addition, risk and impact assessment for low-level HCPs is also outlined, with consideration of clinical information.

A survey of neurological decompression illness in commercial breath-hold divers (Ama) of Japan.

A survey was conducted in the northern district of Yamaguchi, Japan to determine the relationship between neurological diving accidents and risk factors among commercial breath-hold divers (Ama). A questionnaire was distributed to 381 Ama divers who are members of the Ama diving union. We sought information on their dive practices (depth of single dive, single dive time, surface interval, length of dive shifts, lunch break) and the presence or absence of medical problems, such as hypertension, cardiac arrhythmia, diabetic mellitus and other issues. Of the 381 Ama divers, 173 responded (45%): 29 were Funado (assisted-descent using weights) and 144 Cachido (unassisted) divers. Twelve had experienced strokelike symptoms during or after repetitive breath-hold diving; 11 were assisted and one unassisted (Funado vs. Cachido). Only two of 12 divers with neurological diving accidents had musculoskeletal symptoms. Neurological events were significantly correlated with dive depth, dive time, and surface interval; however, they were not related to medical history. Neurological diving accidents are more likely to happen among assisted Ama divers than unassisted ones. Repetitive breath-hold diving with a deep dive depth, long dive time, and short surface interval predisposes divers to decompression illness, which characteristically manifests as cerebral stroke.

[Acute decompression illness following hyperbaric exposure: clinical features of central nervous system involvement].

Decompression illness (DCI) is a general term encompassing all pathological changes secondary to reduction of environmental pressure. This condition has two forms: decompression sickness (DCS) and arterial gas embolism (AGE) secondary to pulmonary barotrauma. Moreover, DCS is categorized as minor, such as limb and/or joint pains or skin rash (Type I), and serious, as in cardiopulmonary and/or central nervous system involvements (Type II). Cerebral and spinal injuries have been symptomatically classified into AGE and DCS. Brain scans of patients with AGE or DCS showed multiple cerebral infarctions in the terminal and/or border zones of the cerebral arteries. Brain involvements of patients in both AGE and DCS show no differences in neurological or neuro-radiological findings. From the neurological and radiological standpoint, it is therefore impossible to distinguish these two conditions. Despite established treatments for neurological DCI (both AGE and DCS), it is unclear whether US Navy treatment Table 6 is preferable to standard hyperbaric oxygen therapy such as 2.4 atmospheres pressure for 90 minutes. Japanese laws and regulations have peculiarities that permit air diving to 90 meters depth, but with explicit prohibition of the use of oxygen for decompression, albeit a limited use of mixed gas is permissible. Moreover, currently the health screening for hyperbaric workers does not include detailed examination of the cardiopulmonary or the central nervous system.

[Central nervous system involvement in patients with decompression illness].

Dysbarism or decompression illness (DCI), a general term applied to all pathological changes secondary to altered environmental pressure, has two forms decompression sickness (DCS) and arterial gas embolism (AGE) after pulmonary barotrauma. Cerebral and spinal disorders have been symptomatically categorized as AGE and DCS, respectively. Magnetic resonance images (MRIs) of divers with DCI showed multiple cerebral infarction in the terminal and border zones of the brain arteries. In addition, there were no differences between MRI findings for compressed air and breath-hold divers. Although the pathogenesis of the brain is not well understood, we propose that arterialized bubbles passing through the lungs and heart involved the brain. From the mechanisms of bubble formation, however, this disorder has been classified as DCS. We propose that there is a difference between clinical and mechanical diagnoses in the criteria of brain DCI. In contrast to brain injury, the spinal cord is involved only in compressed air divers, and is caused by disturbed venous circulation due to bubbles in the epidural space. The best approach to prevent diving accidents is to make known the problems for professional and amateur divers.