Fluorescence cross-correlation spectroscopy (FCCS) is an advanced fluorescence technique that can quantify protein-protein interactions in vivo. Due to the dynamic, heterogeneous nature of the membrane, special considerations must be made to interpret FCCS data accurately. In this study, we describe a method to quantify the oligomerization of membrane proteins tagged with two commonly used fluorescent probes, mCherry (mCH) and enhanced green (eGFP) fluorescent proteins. A mathematical model is described that relates the relative cross-correlation value (f) to the degree of oligomerization. This treatment accounts for mismatch in the confocal volumes, combinatoric effects of using two fluorescent probes, and the presence of non-fluorescent probes. Using this model, we calculate a ladder of f values which can be used to determine the oligomer state of membrane proteins from live-cell experimental data. Additionally, a probabilistic mathematical simulation is described to resolve the affinity of different dimeric and oligomeric protein controls.
Low cost, open-source analytical instrumentation has the potential to increase educational outcomes for students and enable large-scale citizen science projects. Many of these instruments rely on smartphones to collect the data, mainly because they can effectively leverage a dramatic price-to-performance ratio of the optical sensors. However, several hurdles need to be overcome for these devices to be more widely adapted. In this communication we focus on visible spectrophotometers, which are common in chemistry laboratories because of the day-to-day need for quantifying concentration. To make smartphone-based spectrometers practical for wider use, we have designed a 3D-printable spectrophotometer with a dual-beam optical geometry. This geometry allows for sample and reference data to be collected on the same photograph and thus improves the signal-tonoise ratio and reproducibility of the spectra. A universal mounting system was also developed to allow for a wide variety of smartphone form factors to be coupled to the spectrophotometer. To demonstrate potential applications of this device, two assays are reported. The first is a simple illustration of the Beer−Lambert Law with common household dyes. The second is a colorimetric nitrate assay, which shows a quantitative relationship between absorption and nitrate concentration. Kinetic data are also shown for the nitrate assay, which illustrate the long time-stability of the spectral data acquired from the device.
The purpose of this study is to compare the rate of vaginal cuff dehiscence between two different methods of closure in patients undergoing robotic-assisted total laparoscopic hysterectomy and explore variables related to postoperative breakdown. This was a prospective, randomized controlled study with two arms. The control group (Arm 1) underwent single-layer continuous closure while the study group (Arm 2) had three additional imbricating figure-of-X sutures placed in addition to the standard protocol. Of the 263 patients who completed the study, 4 patients (1.49 %) experienced dehiscence of the vaginal cuff. Three of the four patients with dehiscence received the standard single vaginal cuff closure (Arm 1) and the one remaining case of dehiscence underwent the protocol with additional sutures (Arm 2). All patients who experienced dehiscence were current smokers. Our study suggests that there may be benefit in adding additional sutures to the standard single-layer vaginal cuff closure procedure. Physicians should evaluate smoking status before deciding on a vaginal cuff closure method.
An 84-year-old female presented with bilateral scotomas and progressive nyctalopia over 1 year. Best-corrected visual acuity was 20/50 in both eyes with reduced color vision. Goldmann visual field showed bilateral cecocentral scotomas and generalized constriction of the visual fields. This led to an electroretinogram showing an electronegative pattern consistent with autoimmune retinopathies. Infectious workup was negative. Anti-retinal antibodies were positive, leading to a presumed diagnosis of cancer-associated retinopathy (CAR). Imaging showed a previously unknown left renal lower pole mass, and she underwent a radical nephrectomy. Biopsy showed nuclear grade-3 clear cell renal carcinoma staged T1. The patient was treated with oral prednisone with no ocular improvement. We report on a rare case of clear cell renal carcinoma causing CAR. CAR is an important paraneoplastic syndrome to diagnose since the majority of ocular cases precede other manifestations of malignancy. Therefore, a timely diagnosis of CAR can be lifesaving or at least life-extending.
Preseptal cellulitis is an infection of ocular tissue that is often unilateral and caused by extension of sinonasal disease. In rare instances it can lead to life-threatening necrotizing fasciitis. We present here a unique case of bilateral preseptal cellulitis incited by local conjunctivitis caused by prescription eye drops. The patient was immunosuppressed, which allowed her local inflammation to progress to severe infection and, ultimately, to necrotizing fasciitis. This necessitated serial debridement by ophthalmology and otolaryngology teams and a prolonged course of intravenous antibiotics monitored by an infectious disease team. Despite these interventions, the patient's vision did not return to baseline and she had persistent cosmetic and functional deformity. This case is unique due to the inciting incident of new prescription eye drops, the patient's immunosuppressed state leading to severity of infection, and the severe bilateral disease burden.
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