Optical density probes

Several papers have been published in which, instead of concentrating on specific reactions, the technology was highlighted. One, by Marose et al.,7 discusses the various optics, fiber optics, and the probe designs that allow in situ monitoring. They describe the various optical density probes used for biomass determination in situ microscopy, optical biosensors, and specific sensors such as NIR and fluorescence. 

These optical probes are the most universally applicable in situ devices for on-line biomass monitoring up to now [15,16]. Konstaninov et al. [17] tested several absorbance and scattering sensors for real-time biomass concentration monitoring in mammalian cell cultivation processes and Hatch and Veilleux [18] compared optical density probes with oxygen uptake rates, packed cell volume, and off-line cell mass monitoring in commercial fed-batch fermentations of Saccharomyces cerevisiae [19]. In order to minimize influencing effects, special chemometric data treatment is necessary [20]. 

Optical detectors can routinely measure only intensities (proportional to the square of the electric field), whether of optical pulses, CW beams or quasi-CW beams the latter signifying conditions where the pulse train has an interval between pulses which is much shorter than the response time of the detector. It is clear that experiments must be designed in such a way that pump-induced changes in the sample cause changes in the intensify of the probe pulse or beam. It may happen, for example, that the absorjDtion coefficient of the sample is affected by the pump pulse. In other words, due to the pump pulse the transparency of the sample becomes larger or smaller compared with the unperturbed sample. Let us stress that even when the optical density (OD) of the sample is large, let us say OD 1, and the pump-induced change is relatively weak, say 10 , it is the latter that carries positive infonnation. 

When metals have Raman active phonons, optical pump-probe techniques can be applied to study their coherent dynamics. Hase and coworkers observed a periodic oscillation in the reflectivity of Zn and Cd due to the coherent E2g phonons (Fig. 2.17) [56]. The amplitude of the coherent phonons of Zn decreased with raising temperature, in accordance with the photo-induced quasi-particle density n.p, which is proportional to the difference in the electronic temperature before and after the photoexcitation (Fig. 2.17). The result indicated the resonant nature of the ISRS generation of coherent phonons. Under intense (mJ/cm2) photoexcitation, the coherent Eg phonons of Zn exhibited a transient frequency shift similar to that of Bi (Fig. 2.9), which can be understood as the Fano interference [57], A transient frequency shift was aslo observed for the coherent transverse optical (TO) phonon in polycrystalline Zr film, in spite of much weaker photoexcitation [58],... 

A concentrated Ca-D-pantothenate solution (1 mg/ml) is prepared in distilled water and dilutions made as needed. Refrigerated solutions are stable for 6 months. Pantothenate is added at 5, 10, 20, 40, 60, 80, and 100 mpg/ml final concentrations the control flask consists of basal medium alone for estimation of carry-over error—i.e., the pantothenate activity of the inoculum. The details of aseptic technique have been discussed elsewhere (H18, H19). Growth is measured in optical density units with a Welch Densichron, equipped with a red-sensitive probe to minimize blank readings due to the color of the medium. 

Fluorescence probing has also been used by us to investigate the mechanism of vesicle to micelle transformation due to interaction of liposome with sodium dodecyl sulfate micelles. An increase in optical density and hydrodynamic diameter was observed at low SDS concentrations (Fig. 24). The increase was attributed to the incorporation of SDS monomers on liposome vesicles. The point where the hydrodynamic diameter and optical density reached a maximum was proposed to correspond to the saturation of bilayers of the first inflection point. Upon further increase in the SDS concentration... 

There are many ways of testing a gold probe, but the most convincing is by using a known positive sample Therefore, this could be a histological section, an electron microscope specimen, or a dot-blot. Estimation of the concentration of the probe by optical density measurements is a good method to standardize the concentration of probes from one batch to another, but in addition, it is always preferable to test the performance of the probes on known positive samples. 

Attenuated total internal reflection (ATR) probes offer several advantages over other probe types. ATR is a phenomenon that relies on a difference in the index of refraction of a crystal and that of the solution with which it is in contact to prevent light from escaping the crystal. Only the evanescent wave of the light interacts with the solution layer at the crystal face. The result is an optical pathlength of only a few microns. Typical designs make use of faceted crystals or hemispheres (see Figure 6.1). The most common ATR material in the UV-vis is sapphire. In rare cases, fused silica may be used. ATR allows spectra to be taken of neat samples with optical density (OD) of 500-1000... 

The local gas holdup and bubble behavior were measured by a reflective optic fiber probe developed by Wang and co-workers [21,22]. It can be known whether the probe is im-merging in the gas. The rate of the time that probe immerg-ing in the gas and the total sample time is gas holdup. Gas velocity can be got by the time difference that one bubble touch two probes and the distance between two probes. Chord length can be obtained from one bubble velocity and the time that the probe stays in the bubble. Bubble size distribution is got from the probability density of the chord length based on some numerical method. The local liquid velocity in the riser was measured by a backward scattering LDA system (system 9100-8, model TSI). Details have been given by Lin et al. [23]. 

Wu P, Ozturk SS, Blackie JD, Thrift JC, Figueroa C, Naveh D (1995), Evaluation and applications of optical cell density probes in mammalian cell bioreactors, Biotechnol. Bioeng. 45 495-502. 

Fig. 13. Schematic design of the Aquasant probe. This is a sensor for optical density measuring the reflected light. Precision optics focus and collect the incident and the reflected light. Internally, light is guided through optical fibers. Left cross section right front view...

Fig. 14. Schematic design of the Cerex probe. This is a sensor for optical density and mounted vertically in situ. Suspension enters the side drain ports deliberately and can be trapped inside the sensor by powering the solenoid coils the magnetic plunger closes the side ports. In the meantime, the trapped dispersion degasses and bubbles disappear through the upper vent hole. After some time, the optical density reading is declared representative . The next cycle starts with opening the side drain ports...

In order to obtain quantitative results, pump-probe spectra have to be corrected for the optical density of the sample following the Lambert-Beer law of absorption. The correction can be performed in the following way ... 

The optical density (OD) of the sample was varied by repeatedly diluting the mixture with fresh gas until a value of roughly 0.8-1.2 was obtained in some cases, scans were acquired with ODs as low as 0.1. Absorbance measurements were made directly in the cell using a Mattson Research Series FT-IR spectrometer (0.25 cm1 resolution) configured for external beam operation. The optical layout makes it possible to easily switch from making ps pump-probe measurements to recording an IR spectrum. 

One of the most important sensors needed is one that reliably monitors cell density. An IR fiber-optic cell density probe has been used for this because it can directly monitor cell growth (without dilution) in high-cell-density bacterial fermentations. The ability to do an online sample filtration through the use of hollow fibers or rotating filters has made possible continuous, online measurement of glucose, lactate, and other metabolites. However, glucose, nitrogen substrate, and phosphate sensors that can withstand repeated system sterilization are still needed. 

Local voidages for FCC catalyst at various radial positions were measured with an optical fiber probe in a Type A apparatus, from which radial volidage profiles and their probability density functions were computed by Li et al. (1980b), as shown in Figs 20 and 21. When gas velocity is less than the incipient fast fluidization velocity of 1.25 m/s, the radial voidage profile is relatively flat when gas velocity increases further, this profile becomes steeper high in the center. As flow is transformed into pneumatic transport, the... 

Figure 5. Sensitivity of antigen-trapping assay with monoclonal antibodies HSA-1 and HSA-2. HSA-2 (type B antibody) was coated on the plate, and then human albumin (A) or serum (B) was diluted as indicated and added to the wells. Biotin-conjugated HSA-1 served as the specific probe. The optical density at 410 nm (OD410) was measured after 15 min.

In order to illustrate the use of the lactoperoxidase procedure as a vectorial probe, the example to be given is the iodination of the plasma membrane of the human lymphocyte. All manipulations were carried out at 4°. The lymphocytes were isolated and washed free of extracellular protein in isotonic phosphate buffer. The cells were centrifuged at 700 g for 10 min and the supernatant serum was removed. Cells are resuspended in 5 to 10 volumes of phosphate buffer and centrifuged free of the supernatant. This procedure was repeated four times or more until the supernatant gave a 280 nm reading below an optical density of 0.1 absorbance. The pellet of washed cells were then suspended in 3 volumes of isotonic phosphate buffer to produce 2 x 10 cells/ml. One millicurie of I was added and 20 nmol of lactoperoxidase for each milliliter of cell suspension. Ten microliter portions of 1 mM peroxide was added at 1 min intervals. The lymphocytes were then spun down at 500 g for 5 min and washed three times with phosphate-buffered saline. 

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