Introduction
Think of your old manual Spectronic 20, or your direct reading spectrophotometer that you use in your lab. You line up your samples in a row. In front of them, you place some small sample cups or maybe even a series of cuvettes, and you pipette a known whole of sample into each cup. You then add a reagent and somehow mix the reagent and sample. You do this for each sample. You may have more reagents to add so you repeat the whole process until all reagents are added. Then you start a timer. When the timer beeps you know you have a safe bet "time window" to read the absorbance (or concentration) of your samples. You read by manually transferring the color-developed sample to a spectrometer cuvette, by using a peristaltic pump to change the sample to a flow cell already in the spectrometer, or by inserting the tube or cuvette that you used to invent the sample color in. Then, you press a button to send the reading to a printer, a computer program, or you manually narrative the reading onto a laboratory worksheet.
Did you shake and mix every sample exactly the same way every time? Will you mix them the same way every day? Will every interpreter run them exactly the same way you have?
Is there color or turbidity in the samples? Should you zero your instrument with each sample, or only with reagent water blanks?
Is the exact time you read the final absorbance critical?
The process described is what you are automating by using a assorted analyzer. Instead of lining up samples, you are pouring aliquots into sample cups that are settled on an auto sampler tray. Instead of transferring a known whole of sample to a cuvette, the assorted analyzer does. Instead of adding reagents and mixing, the assorted analyzer does. Instead of beginning a timer, the assorted analyzer does. Instead of reading the absorbance, recording the reading, and calculating a result the assorted analyzer does.
The analyzer has automated approximately all the simple colorimetric methods for you. Sample volume is measured and dispensed exactly the same way, every time. Reagents are added and mixed exactly the same way every time. The timer is set and absorbance is measured exactly the same way every time. Results are calculated exactly the same way every time.
The assorted analyzer pipettes, dilutes, adds reagents, mixes, calibrates, measures, calculates, and reports all for you. You pick a formula by keyboard. There is no hardware to manually change, no cartridge to rinse out, no baselines to monitor, no wavelength filters to change. Sample and reagent volumes are carefully by a selection in a computer program, not by the internal diameter of a peristaltic pump tube.
The assorted analyzer has done a lot for you but it cannot control nor do everything. It cannot accurately get ready the stock calibration standard for you, even though it can accurately dilute it. It cannot certify the standards and samples were settled on the auto sampler tray in the right order. It cannot get ready the reagents for you or certify they were settled in the right order; however, it can monitor their lucidity and remind you where they are supposed to go. It cannot make sure you've entered the proper sample Id for each sample position, however, it can certify that the result obtained for that sample position is traceable to the Id you entered. It cannot know the sample lot Id for each standard or reagent, but if you enter those Id's into the software, it can certify traceability of those reagents with your sample sets.
The software and built in electronics permanently monitor and adjust lamp voltage so that absorbance readings do not drift. Drift is coarse in flow analyzers because the peristaltic pump tubing delivers reagents by proportion. The assorted analyzer delivers the exact whole of sample and reagent every time. These volumes do not change. The assorted analyzer has a fixed path distance if the assorted analyzer does not change color-developed sample to an additional one cuvette, or flow cell, for measurement. In addition, if, the assorted analyzer reads straight through the walls of the cuvette the calibration curve is normally more carport and or reproducible than your reagents and standards.
Change your thoughts on calibration
Beer's law states that the absorbance is equal to the absorbtivity times the path distance times the concentration. It seems, however, sometimes we do not believe that Beer's law is a law. I say this because agreeing to this law, the absorbtivity is a constant. When the path distance is fixed (always the same), the path distance is a constant as well development the only changeable the concentration. Therefore, you get ready standards of a known concentration, quantum the absorbance and conclude the absorbtivity. Assuming you can get ready reagents exactly the same way every time, quantum the same volume every time, and incubate your samples the same whole of time every time, there should be no surmise to assume that the absorbtivity would change. If the absorbtivity does not change, then there is no surmise to calibrate every day. Moreover, if the absorbtivity is not changing, you could in fact be introducing error every time you calibrate because you may not be taking into list random errors that occur in the middle of analysts or even with yourself as you inadvertently vary your technique on a day-to-day basis.
As mentioned previously, daily calibration is required for continuous flow methods because flow methods proportion the reagents and sample using a peristaltic pump. Those pump tubes are changing with time changing the relative proportion of sample and reagents. Flow analyzers are still incredibly accurate, it is just you need to calibrate each time.
Calibrating consumes time. Especially exact ones where you took great care to ensure your standards and reagents are fresh.
A manual spectrometer does not necessarily require a calibration each time. Many methods written for manual spectrometers merely say, "analyze a check standard with each sample set". In fact, the stability of the calibration curve is the underlying opinion behind direct reading spectrophotometers and filter wheel methods. For many colorimetric tests, the stability of the curve far exceeds the stability of the standards or the reagents. Some examples are nitrite and phosphate.
A assorted analyzer should not require daily calibrations and should allow us to extrapolate more the ion chromatography, gas chromatography, and manual direct reading spectrometer opinion of the chronic Calibration Verification, or Ccv. As mentioned, the surmise the assorted analyzer curves are carport is that the robot exactly reproduces everything every time. You cannot do this because you are not a robot, the assorted analyzer, however, is.
A manual formula uses more reagent and sample volume because we, as humans, cannot work in fact with small volumes. A flow system uses more reagent than a assorted analyzer because a flow instrument is continuously pumping reagent straight through the system.
Discrete analyzers that quantum the sample absorbance within the same holder that the reaction occurred originate less waste than instruments that wash the vessel, or use a flow cell. In fact, adequately rinsing a flow cell requires valuable rinsing in the middle of samples development the waste volume generated essentially equivalent to that of a micro-flow Segmented Flow Analyzer, or Low Flow Flow Injection Analyzer.
The assorted analyzer uses significantly less reagent, and generates significantly less waste than manual methods. This chart illustrates an unscaled down manual formula using the exact volumes described in standard Methods. The waste generated for the manual formula does not take into list washing of glassware. As mentioned earlier, an analyzer that washes cuvettes or rinses a flow cell will originate more waste than indicated here.
Eliminate the possibility of contamination, or false positives
The assorted analyzer measuring the absorbance of a color reacted sample contained in personel cuvettes. Unlike flow analysis, there is no possibility of interaction in the middle of samples and unlike flow analysis; the user can visually observe the reaction product during and after analysis.
Using a assorted analyzer, the interpreter can observe the reaction during color improvement and after the test is complete. The interpreter can take off the reaction segments and verify that dispensed volumes are repeatable, that there are no bubbles or turbidity, and that the color looks correct. A flow analyzer does not give the interpreter the quality to visually observe and qualitatively certify the accuracy of his or her results.
A assorted analyzer dispenses, reacts, incubates, and measures all within the reaction cuvette without transferring to a flow cell. Analyzers that change to a flow cell are not "true" assorted analyzers, but instead, are hybrids in the middle of flow and discrete. The hybridization is done to accomplish lower detection limits; however, the benefit of the individually contained reaction and absence of carryover is lost. In addition, since these analyzers require as much rinse as a flow analyzer to take off preceding samples, waste generation is as high as flow. Given this, and the increased possibility of environmental contamination or analyte loss that occurs from open-air heated reactions, you may as well have a flow analyzer.
Chemical reactions occur in individually contained segments
All assorted analyzers have reaction segments. Some analyzers do chemical reactions in a cuvette segment and then change the reacted sample to a flow cell. This type of analyzer is a hybrid of assorted and flow, and not a true assorted analyzer. A true assorted analyzer reacts and measures the sample within the visual cuvette. Some analyzers wash the visual cuvette in the middle of tests. Washing in the middle of tests enables more samples to be analyzed per cuvette; however, the washing cannot certify that there is no residual contamination that remaining after the washing process. Other assorted analyzers utilize disposable visual quality cuvettes.
Washing in the middle of tests enables more samples to be analyzed per cuvette; however, the washing cannot certify that there is no residual contamination not thoroughly removed by the washing process. This residual contamination can come from preceding samples, or more likely, from the reagents used in processing the preceding samples. The built in computerized checking of visual quality cannot verify absence of chemical contamination.
Analyzers that use a flow cell still react samples in some sort of cuvette. It is the whole of reaction vessels on the assorted analyzer that limit the whole of tests that the assorted can run in a singular walk away operation. If the assorted analyzer has 100 sample positions and 200 reaction cuvettes, then the analyzer can run 100 samples for 2 tests each. The assorted analyzer with the flow cell must rinse the flow cell in the middle of each sample, and rinse vigorously in the middle of each test. Think that a two-channel flow analyzer can analyze 100 samples for two tests each in less than half the time as a assorted analyzer with a flow cell. Also, Think that the flow analyzer generates no more waste than the assorted analyzer with a flow cell. If the required testing is a lot of samples for one or two tests it makes more sense to use a flow analyzer.
Reagents can interfere as cross contamination in the middle of samples. Using disposable personel reaction cuvettes thoroughly eliminates the possibility of contamination. For instance, the cadmium discount nitrate test contains valuable amounts of ammonia in the buffer reagent and phosphate in the color reagent. Using personel disposable cuvettes ensures that there is no contamination. Washing cuvettes, or using a flow cell, means you can never be sure.
Using disposable visual cuvettes is the only way you can certify no carryover in the middle of tests or samples. The opinion is similar to use of disposable petri dishes, disposable pipette tips, and disposable hypodermic needles. The assorted analyzer in fact and rapidly analyzes many tests on singular sample solutions. Only disposable individually contained reactions ensure that there is no interaction in the middle of samples or tests.
Let the robot do your pipetting.
When you manually pipette samples you, hopefully, use a separate pipette per sample. If not, you will at least rinse it in in the middle of samples, and perhaps with sample prior to transferring your sample aliquot to the sample container. This is to avoid carryover in the middle of samples. A flow analyzer uses an auto sampler. The sampling probe immerses in the wash hub rinsing the exterior of the probe, and pulls wash explication from the hub and into the analytical cartridge.
A assorted analyzer also uses a probe; however, it operates differently than flow analyzers. A assorted analyzer's level detect mechanism ensures that the probe immerses into the sample or reagents no added than valuable to withdraw the required sample aliquot. The probe then washes itself on the exterior at the wash hub and pushes the sample or reagent out into the sample cuvette. in the middle of dispenses, the probe pushes excess wash water out ensuring no carryover. In other words, unlike a flow system that only pulls sample in one direction, the sampling probe on a assorted analyzer is bidirectional pulling reagent and sample into its internal tubing only far sufficient to withdraw the exact volume and then dispensing it by pushing it out the other way.
The engine can think.
When doing a manual test you know if you ran out of reagent or sample. A flow analyzer does not know. A flow analyzer could end up aspirating from empty sample cups or empty reagent bottles all night long and think it is still running samples. A assorted analyzer with level detection prevents this. The level detect mechanism is a capacitance detector that senses the incompatibility in the middle of liquid and air. The assorted software calculates the volume of reagents and samples based on the height of liquid. The software continuously monitors sample and reagent volumes and will not continue the test when it detects that reagents or samples have "run out".
The sampling depth on a flow analyzer is normally adjustable by the user and is normally towards the bottom of the sample vial. On a assorted analyzer, the depth the probe immerses in a sample explication is a result of programming or instrument design. The depth sampled on the Oi assorted analyzer is carefully by the level detect mechanism and the sample aliquot required for the test. For instance, if 200 micro liters is required the probe will immerse just below 200 micro liters as carefully by the volume of the cup and the liquid level detected and withdraw a software-defined whole above 200 micro liters. In other words, the assorted analyzer samples from the top 300 micro liters of sample solution. The probe only immerses as far as it has to. This minimizes potential carryover contamination, and speeds the process. In this way dispensing and rinsing is fast and there is no sample or reagent carried to an additional one on the sides of the probe.
When sampling from the top of the sample cup there is a risk of loss of a volatile analyte from the top of the explication or the risk of the adsorption of an analyte from the laboratory air into the top of the solution. For instance, trace cyanide in near neutral explication can be slowly lost from the top layer of sample explication into the lab air. This is especially clear with lower concentrations such as 10 ppb.
Gain of the analyte is potential as well. Ammonia is a coarse laboratory contaminant. Ammonia easily adsorbs into acidified solutions. It is potential for ammonia to be "pulled" from laboratory air into the sample solution. A flow analyzer would not as easily detect this loss or gain because it samples from the bottom of the sample cup.
There are some drawbacks
A assorted analyzer reacts sample in a heated cup that is open to allow the probe to dispense samples and reagents. The heat increases reaction rates and is especially foremost for chemistries such as ammonia that are slow to invent color. In manual testing the reagents are added in open containers, however, the holder shape can vary and the holder can be capped during mixing, heating, and color reaction. When flow analyzers were first introduced one of the key advantages that gained its acceptance over manual methods was that reactions occurred enclosed within the tubing limiting its exposure to laboratory air. In this aspect, assorted analyzers are kind of a step backwards.
There are valuable advantages.
Similar to holding a color developing reaction in its own holder till it reaches a color maximum, assorted analyzers can also hold intermediate reactions for long periods of time without risk of carryover, dilution into a carrier reagents, or excessive dispersion. This can be especially useful in enzyme or discount reactions where reaction rates are slow. A flow analyzer would require long delay coils resulting in very complex Sfa chemistry manifolds. Often elevated climatic characteristic is used to speed reactions, but in some chemistry, there are limits to the maximum temperatures possible. Since assorted analyzer reactions are occurring in individually contained cuvettes, the time delay in the middle of reagent additions on assorted analyzers is petite only by software. This is a valuable benefit over flow chemistry.
In manual methods, obviously, the operator prepares all the calibration standards from a stock solution, dilutes any Qc samples from a stock solution, dilutes samples known to be over calibration prior to color development, and dilutes samples that were over calibration once he or she notices that they are. Unless you have an added auto-dilutor attached to your flow analyzer, you will still be diluting standards and over calibration samples. Auto-dilution is an integral function of a assorted analyzer. The dilutions can be preset during sample table entry if you know that the samples need to be diluted. Methods can be programmed such that they dilute every sample and standard all the time, or the instrument can be programmed so that over calibration, samples are diluted and re analyzed.
An interpreter changes a manual or flow formula from one to the next by memory, or by referring to the Sop. How well this singular interpreter performs the procedure is dependent upon his mood, the time of day, his experience with the method, the availability of equipment, and many other unquantifiable variables. It is potential to acquire good results and bad results by the same manually performed method. A flow analyzer analyzes everything the same every time assuming it is set up the same every time. This assumption is valid with experienced flow determination technicians; however, if the technician does not understand flow or if there are many users results will vary. Farranging training and documentation is valuable to certify that results conform to good automated lab practices.
The assorted analyzer formula is selected by mouse click when scheduling analyses on the sample tray. The formula conditions do not change. In fact, assuming you have accurately calibrated your formula the calibration is stored within the method. This means that an untrained interpreter that only knows what buttons to press is able to acquire identical results to even the most experienced analyst.
Most analytes performed in an environmental yielding laboratory cannot be bench spiked. If the analyte requires a introductory distillation, digestion, or discharge the spiking is done prior to the introductory sample process. I comprehend that many labs do not distill ammonia or Fluoride and I would argue that if you are reporting yielding testing for the clean water act you would good seriously Think changing your Sop. Other parameters that can't be spiked are those that are too high to spike within the matrix without introductory dilution, such as Ca, Mg, Cl, So4, and analytes like alkalinity that just are not spiked.
This shortens the list of potential analytes for the automated spiking function to nitrite, phosphate, Sulfide, Chromium Vi, and some others. On these, I defer back to the previous slide and ask if the potential error is worth the risk for so few tests.
Summary
Benefits of assorted analyzers consist of decreased reagent consumption, decreased waste generated, and ease of use among other things. The most valuable benefit of the assorted analyzer, however, is that it can eliminate the traditional opinion of habit determination and allow you to run samples as you receive them instead of storing them until there is sufficient sitting around to make a flow or Ic determination worthwhile. If you take benefit of the calibration stability of the assorted analyzer, and accurately get ready a calibration that can then be used by approximately any interpreter in subsequent uses an added benefit is that the results are the same regardless of who uses the machine.
Think of those short holding time samples. The phosphate, the nitrites, the chromium Vi, and residual chlorine. These analytes cause the environmental lab to stop everything just to get the determination done on time. Think of the other analytes that come in periodically, but maybe not frequently. perhaps silica, ferrous iron and sulfide. How do you certify these tests followed the Sop? Instead of mental of the assorted analyzer as something to replace a flow instrument, think of it as something to supplement a flow instrument. If you have hundreds of samples for one or two tests routinely and for the same analyte you are not going to save money by switching these tests to a assorted analyzer. Where you will save money and great endeavor is removing unnecessary strain from the flow analyzer and your analysts by performing the non - habit or "rush" tests on a assorted analyzer. It is potential for the sample login man to analyze samples as received for approximately every colorimetric test that does not require a digestion. In other words, as soon as the sample is logged in it could be immediately run for nitrite, phosphate, chromium Vi, nitrate, ammonia, chloride, and sulfate. In this example, instead of putting samples in a refrigerator to be gathered for determination at a later time, they end up being run by ice chest and by client as soon as they are received.
If everything is to run on the assorted analyzer, then acquire your samples in a vial that fits on the assorted analyzer. You no longer need to change liquid from holder A to auto sampler vial B, the sample bottle can be the auto sampler vial. Not only does this save time, but it saves shipping as well. Instead of large ice chests, you use tiny mailers.
To summarize, the true benefit of a assorted analyzer is that its built in features allow any interpreter to get the same results every time. assorted analyzers are very simple to use requiring minimal software training. Once set up for your laboratory, properly applied methods allow you to modify your daily routines and analyze samples as soon as they come in. Either you are an environmental lab, research, process control, or municipality assorted analyzers can be used effectively in your operation. Currently, the full power of assorted analyzers is petite by tradition and by regulation. Once we start to invent methods for assorted analyzers instead of using assorted analyzers to run methods developed for flow we will be able to see greater throughput, less variability, and lower Mdl.
