AutoAnalyzer

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The AutoAnalyzer is an automated analyzer using a flow technique called continuous flow
analysis (CFA), first made by the Technicon Corporation. The instrument was invented 1957
by Leonard Skeggs, PhD and commercialized by Jack Whitehead's Technicon Corporation.
The first applications were for clinical analysis, but methods for industrial analysis soon
followed. The design is based on separating a continuously flowing stream with air bubbles.

Contents
1 Operating principle

o 1.1 Flow injection analyzer

o 1.2 Dialyzer module

o 1.3 Recording of results

2 Commercialization

3 Clinical analysis

4 Industrial analysis

5 Current Uses

6 Method sheets

7 Notes

8 External links

Operating principle
In continuous flow analysis (CFA) a continuous stream of material is divided by air bubbles
into discrete segments in which chemical reactions occur. The continuous stream of liquid
samples and reagents are combined and transported in tubing and mixing coils. The tubing
passes the samples from one apparatus to the other with each apparatus performing different
functions, such as distillation, dialysis, extraction, ion exchange, heating, incubation, and
subsequent recording of a signal. An essential principle of the system is the introduction of air
bubbles. The air bubbles segment each sample into discrete packets and act as a barrier
between packets to prevent cross contamination as they travel down the length of the tubing.
The air bubbles also assist mixing by creating turbulent flow (bolus flow), and provide
operators with a quick and easy check of the flow characteristics of the liquid. Samples and
standards are treated in an exactly identical manner as they travel the length of the tubing,
eliminating the necessity of a steady state signal, however, since the presence of bubbles
create an almost square wave profile, bringing the system to steady state does not
significantly decrease throughput ( third generation CFA analyzers average 90 or more
samples per hour) and is desirable in that steady state signals (chemical equilibrium) are more
accurate and reproducible.[1]

A continuous flow analyzer (CFA) consists of different modules including a sampler, pump,
mixing coils, optional sample treatments (dialysis, distillation, heating, etc.), a detector, and
data generator. Most continuous flow analyzers depend on color reactions using a flow
through photometer, however, also methods have been developed that use ISE, flame
photometry, ICAP, fluorometry, and so forth.

Flow injection analyzer

Main article: Flow Injection Analyzer

Flow injection analysis (FIA), was introduced in 1975 by Ruzicka and Hansen,[2] The first
generation of FIA technology, termed flow injection (FI), was inspired by the AutoAnalyzer
technique invented by Skeggs in early 1950s.[citation needed] While Skeggs' AutoAnalyzer uses air
segmentation to separate a flowing stream into numerous discrete segments to establish a
long train of individual samples moving through a flow channel, FIA systems separate each
sample from subsequent sample with a carrier reagent. While the AutoAnalyzer mixes sample
homogeneously with reagents, in all FIA techniques sample and reagents are merged to form
a concentration gradient that yields analysis results.

FIA methods can be used for both fast reactions as well as slow reactions. For slow reactions,
a heater is often utilized. The reaction does not need to reach completion since all samples
and standards are given the same period to react. For typical assays commonly measured with
FIA (e.g., nitrite, nitrate, ammonia, phosphate) it is not uncommon to have a throughput of
60-120 samples per hour.

FIA methods are limited by the amount of time necessary to obtain a measurable signal since
travel time through the tubing tends to broaden peaks to the point where samples can merge
with each other. As a general rule, FIA methods should not be used if an adequate signal
cannot be obtained within two minutes, and preferably less than one.[citation needed] Reactions that
need longer reaction times should be segmented. However,considering the number of FIA
publications and wide variety of uses of FIA for serial assays, the "one minute" time
limitation does not seem to be a serious limitation for most real life assays.[citation needed] Yet,
assays based on slow chemical reactions have to be carried either in stopped flow mode
( SIA) or by segmenting the flow.

OI Analytical, in its gas diffusion amperometric total cyanide method, uses a segmented flow
injection analysis technique that allows reaction times of up to 10 minutes by flow injection
analysis.[3]

Technicon experimented with FIA long before it was championed by Ruzicka and Hansen.
Andres Ferrari reported that analysis was possible without bubbles if flow rates were
increased and tubing diameters decreased.[4] In fact, Skegg's first attempts at the auto analyzer
did not segment. Technicon chose to not pursue FIA because it increased reagent
consumption and the cost of analysis.[citation needed]

The second generation of the FIA technique, called sequential injection analysis (SIA), was
conceived in 1990 by Ruzicka and Marshal, and has been further developed and miniaturized
over the course of the following decade.[citation needed] It uses flow programming instead of the
continuous flow regime (as used by CFA and FIA), that allows the flow rate and flow
direction to be tailored to the need of individual steps of analytical protocol. Reactants are
mixed by flow reversals and a measurement is carried out while the reaction mixture is
arrested within the detector by stepping the flow. Microminiaturized chromatography is
carried out on microcolumns that are automatically renewed by microfluidic manipulations.
The discrete pumping and metering of microliter sample and reagent volumes used in SI only
generates waste per each sample injection. The enormous volume of FI and SI literature
documents the versatility of FI and SI and their usefulness for routine assays (in soil, water,
environmental, biochemical and biotechnological assays) has demonstrated their potential to
be used as a versatile research tool.

Dialyzer module

In medical testing applications and industrial samples with high concentrations or interfering
material, there is often a dialyzer module in the instrument in which the analyte permeates
through a dialysis membrane into a separate flow path going on to further analysis. The
purpose of a dialyzer is to separate the analyte from interfering substances such as protein,
whose large molecules do not go through the dialysis membrane but go to a separate waste
stream. The reagents, sample and reagent volumes, flow rates, and other aspects of the
instrument analysis depend on which analyte is being measured. The autoanalyzer is also a
very small machine

Recording of results

Previously a chart recorder and more recently a data logger or personal computer records the
detector output as a function of time so that each sample output appears as a peak whose
height depends on the analyte level in the sample.

Commercialization
Technicon sold its business to Revlon in 1980 [5] who later sold the company to separate
clinical (Bayer) and industrial (Bran+Luebbe - now SEAL Analytical) buyers in 1987. At the
time, industrial applications accounted for about 20% of CFA machines sold.
In 1974 Ruzicka and Hansen carried out in Denmark and in Brasil initial experiments on a
competitive technique, that they termed flow injection analysis (FIA). Since then the
technique found world wide use in research and routine applications, and was further
modified through miniaturization and by replacing continuous flow with computer controlled
programmable flow.

During the 1960s industrial laboratories were hesitant to use the autoanalyzer. Acceptance by
regulatory agencies eventually came about by demonstration that the techniques are no
different from a recording spectrophotometer with reagents and samples added at the exact
chemical ratios as traditionally accepted manual methods.[6]

The best known of Technicon's CFA instruments are the AutoAnalyzer II (introduced 1970),
the Sequential Multiple Analyzer (SMA, 1969), and the Sequential Multiple Analyzer with
Computer (SMAC, 1974). The Autoanalyzer II (AAII) is the instrument that most EPA
methods were written on and reference.[citation needed] The AAII is a second generation segmented
flow analyzer that uses 2 millimeter ID glass tubing and pumps reagent at flow rates of 2 - 3
milliliters per minute. Typical sample throughput for the AAII is 30 - 60 samples per hour.[7]
Third generation segmented flow analyzers were proposed in the literature,[8] but not
developed commercially until Alpkem introduced the RFA 300 in 1984. The RFA 300 pumps
at flow rates less than 1 milliliter per minute through 1 millimeter ID glass mixing coils.
Throughput on the RFA can approach 360 samples per hour, but averages closer to 90
samples per hour on most environmental tests. In 1986, Technicon (Bran+Luebbe) introduced
its own microflow TRAACS-800 system.[9]

Bran+Luebbe continued to manufacture the AutoAnalyzer II and TRAACS, a micro-flow

analyzer for environmental and other samples, introduced the AutoAnalyzer 3 in 1997 and the
QuAAtro in 2004. The Bran+Luebbe CFA business was bought by SEAL Analytical in 2006
and they continue to manufacture, sell and support the AutoAnalyzer II/3 and QuAAtro CFA
systems, as well as Discrete Analyzers.

And there are other manufacturers of CFA instruments.

Skalar Inc., subsidiary of Skalar Analytical, founded in 1965, which has its head office in
Buford, is since its founding an independent company, fully owned by its personnel.
Development in robotic analyzers, TOC and TN equipment, and monitors has extended the
product lines of its long life SAN++ Continuous Flow Analyzers. Software packages for data
acquisition and analyzer control are also in house products, running with latest software
demands and handles all analyzer hardware combinations.

Astoria-Pacific International, for example, was founded in 1990 by Raymond Pavitt, who
previously owned Alpkem. Based in Clackamas, Oregon, U.S.A., Astoria-Pacific
manufactures its own micro-flow systems. Its products include the Astoria Analyzer lines for
Environmental and Industrial applications; the SPOTCHECK Analyzer for Neonatal
screening; and FASPac (Flow Analysis Software Package) for data acquisition and computer
interface.

FIAlab Instruments, Inc., in Bellevue Washington, also manufactures several analyzer

systems.
Alpkem was purchased by Perstorp, and then later by OI Analytical in College Station Texas.
OI Analytical manufactures the only segmented flow analyzer that uses polymeric tubing in
place of glass mixing coils. OI is also the only major instrument manufacturer that provides
segmented flow analysis (SFA) and flow injection analysis (FIA) options on the same
platform.

Clinical analysis
AutoAnalyzers were used mainly for routine repetitive medical laboratory analyses, but they
had been replaced during the last years more and more by discrete working systems which
allow lower reagent consumption. These instruments typically determine levels of albumin,
alkaline phosphatase, aspartate transaminase (AST), blood urea nitrogen, bilirubin, calcium,
cholesterol, creatinine, glucose, inorganic phosphorus, proteins, and uric acid in blood serum
or other bodily samples. AutoAnalyzers automate repetitive sample analysis steps which
would otherwise be done manually by a technician, for such medical tests as the ones
mentioned previously. This way, an AutoAnalyzer can analyze hundreds of samples every
day with one operating technician. Early AutoAnalyzer instruments each tested multiple
samples sequentially for individual analytes. Later model AutoAnalyzers such as the SMAC
tested for multiple analytes simultaneously in the samples.

In 1959 a competitive system of analysis was introduced by Hans Baruch of Research

Specialties Company. That system became known as Discrete Sample Analysis and was
represented by an instrument known as the "Robot Chemist." Over the years the Discrete
Sample Analysis method slowly replaced the Continuous Flow system in the clinical
laboratory.[10]

Industrial analysis
The first industrial applications - mainly for water, soil extracts and fertilizer - used the same
hardware and techniques as clinical methods, but from the mid-1970s special techniques and
modules were developed so that by 1990 it was possible to perform solvent extraction,
distillation, on-line filtration and UV digestion in the continuously flowing stream. In 2005
about two thirds of systems sold worldwide were for water analysis of all kinds,[citation needed]
ranging from sub-ppb levels of nutrients in seawater to much higher levels in waste water;
other common applications are for soil, plant, tobacco, food, fertilizer and wine analysis.

Current Uses
AutoAnalyzers are still used for a few clinical applications such as neonatal screening or
Anti-D, but the majority of instruments are now used for industrial and environmental work.
Standardized methods published by the ASTM (ASTM International), the US Environmental
Protection Agency (EPA) as well as the International Organization for Standardization (ISO)
for environmental analytes such as nitrite, nitrate, ammonia, cyanide, and phenol.
Autoanalyzers are also commonly used in soil testing laboratories, fertilizer analysis, process
control, seawater analysis, air contaminants, and tobacco leaf analysis.

Method sheets
Main article: Colorimetric analysis

Technicon published method sheets for a wide range of analyses and a few of these are listed
below. These methods and later methods are available from SEAL Analytical. Method lists
for manufacturers instruments are readily available on their websites.

Sheet no. Determination Sample Main reagent(s) Colorimeter

N-1c Urea nitrogen Blood or urine Diacetyl monoxime 520 nm
N-2b Glucose Blood Potassium ferricyanide 420 nm
N-3b Kjeldahl nitrogen Foodstuffs Phenol & hypochlorite 630 nm
P-3b Phosphate Boiler water Molybdate 650 nm

Notes
1. Coakly, William A., Handbook of Automated Analysis, Mercel Dekker, 1981 p
61

2. J., Rulika; Hansen, E. H., (1975). "Flow injection analyses: I. New concept of
fast continuous-flow analysis". Anal. Chim. Acta 78: 145157. doi:10.1016/S0003-
2670(01)84761-9.

3. http://www.oico.com/default.aspx?id=product&productid=94

4. (Technicon Symposia, 1967, Vol I)

5. Whitehead Institute - Making clinical progress

6. Coakly, William A., Handbook of Automated Analysis, Marcel Dekker, Inc.,

1981

7. Ewing, Galen Wood, Analytical Instrumentation Handbook, Second Edition

pp152

8. C.J. Patton, PhD. Dissertation, Michigan State University (1982)

9. Ewing, Galen Wood, Analytical Instrumentation Handbook, Second Edition

pp153

10. Rosenfeld, Louis. Four Centuries of Clinical Chemistry. Gordon and Breach
Science Publishers, 1999. ISBN 90-5699-645-2. Pp. 490-492