n engl j med 368;24 nejm.org june 13, 2013 2319s o u n d i.docx

n engl j med 368;24 nejm.org june 13, 2013 2319
s o u n d i n g b o a r d
T h e n e w e n g l a n d j o u r n a l o f m e d i c i n e
How Point-of-Care Testing Could Drive Innovation
in Global Health
Ilesh V. Jani, M.D., Ph.D., and Trevor F. Peter, Ph.D., M.P.H.
The investment in health services in low- and mid-
dle-income countries has increased substantially
in recent years.1 Such investment has been led by
unprecedented efforts to combat major diseases,
enabled by the availability of lower-cost and effec-
tive drug regimens for treatment and prophylaxis,
along with improved vector control. As health
services have expanded, so has the demand for
diagnostic tests that are essential in identifying
patients, determining prognosis, monitoring treat-
ment, and assessing the efficacy of prevention.2
Classic diagnostic technologies are not well
suited to meeting the expanded testing needs.
Laboratory tests require complex infrastructure,
skilled technicians, and a stable supply of elec-
tricity, all of which are scarce, particularly in
nonurban areas. Traditional testing is usually
performed in remote laboratories, which increas-
es the cost and inconvenience of accessing health
care and leads to a high number of patients who

leave the system before a diagnosis is established.3
These limitations are a critical barrier to equity
in health services. Microscopy requires less in-
frastructure and is more widely available, but it
can be inaccurate (e.g., sputum tests for tubercu-
losis) or slow and underutilized (e.g., smear tests
for malaria, schistosomiasis, and other parasitic
infections).4-6 Many patients with tuberculosis or
malaria are simply treated on the basis of a pre-
sumptive clinical diagnosis. Although convention-
al laboratory testing and microscopy will still be
needed, it is expected that faster and more ac-
curate point-of-care diagnostic tests that do not
require laboratory infrastructure will play an in-
creasing role in expanding health care in low- and
middle-income countries.7
T h e S h if t t o w a r d P o in t- o f - C a r e
T e s t in g
Rapid point-of-care testing for diabetes, anemia,
pregnancy, human immunodeficiency virus (HIV),
and malaria have long been available and have be-
come common diagnostic tools in both high- and
low-income countries (Fig. 1). The first generation
of point-of-care testing relied on easy-to-detect
biomarkers, such as antibodies, antigens, and sim-
ple biochemical reactions. Such biomarkers are
also increasingly used in point-of-care tests for a
wide range of infectious diseases (e.g., syphilis,
hepatitis, measles, schistosomiasis, and tricho-
moniasis) and for applications such as blood
typing.8-11
A second generation of point-of-care diagnos-

tics is now on the horizon, partly because of re-
cent industry and donor investment. These tests
detect more complex and less accessible biomark-
ers, such as nucleic acids and cell-surface markers,
and take advantage of advances in microfluidics,
microelectronics, optical systems, and laboratory-
on-a-chip nucleic acid test (NAT)–based amplifi-
cation and detection techniques.12,13 The first
applications of these technologies have included
enumeration of CD4+ T cells, NAT-based diagno-
sis of tuberculosis and drug-resistance screening,
and testing of HIV viral load.14-16 Additional ap-
plications are in the pipeline for other blood-
borne and respiratory infections.
A third generation of technologies will enable
simultaneous detection of multiple targets (multi-
plexing) and will use more accurate biomarkers.
Advances in engineering and test chemistry will
produce devices that are smaller, simpler to op-
erate, and potentially instrument-free,17 enabling
reliable home-based testing or self-testing. These
technologies will extend a wider range of diag-
nostics from the laboratory into clinics and com-
munities.
Point-of-care testing can have a transforma-
tive effect on health care. Rapid HIV tests cata-
lyzed increased rates of case finding that have
driven global efforts in HIV prevention and treat-
ment.18 Malaria rapid tests have been instrumen-
tal in raising testing rates in Africa for suspected
The New England Journal of Medicine
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n engl j med 368;24 nejm.org june 13, 20132320
Detection targets
Antibodies
Antigens
Simple biochemical reactions
Typical samples
Test cartridges
Sample (e.g., capillary blood,
oral fluid, or urine) is inserted
into disposable test cartridge
Samples
Capillary blood, oral fluid,
urine, breath, and other
samples
Multiple test formats
Handheld lab-on-a-chip devices
Disposable tests (no instruments)
Doctor’s office desk-based devices
Devices will fully automate
testing and analysis or
display of results

Transmission of results
Devices are likely to have wireless
connectivity to transmit result data
Small instruments process and read results
Common test formats
First generation of POC diagnostic testing
Examples
Rapid test strips and dipsticks (HIV
antibody and antigen, malaria antigen,
urine biochemistry, and pregnancy tests)
Simple instruments (glucometers and
hemoglobin meters)
Detection targets
Whole cells
DNA or RNA using PCR or other
nucleic acid detection method
Examples
CD4-cell count
HIV viral load
Tuberculosis diagnosis and potential
drug resistance
Potential detection targets
Nucleic acid sequencing
Advanced protein analysis (proteomics)
Examples
Antiviral and antibiotic drug-resistance
screening

Differential diagnosis (e.g., viral rash and
fever, childhood diseases, antenatal tests)
Home-based self-testing
A
Second generation of POC diagnostic testingB
Next generation of POC diagnostic testingC
Urine
Automated
reading
Manually read cartridge-
based strips
Manually read
dipsticks
Oral
fluid
Capillary blood
Lateral-flow test
Vertical-flow test
05/29/13
AUTHOR PLEASE NOTE:
Figure has been redrawn and type has been reset
Please check carefully

Author
Fig #
Title
ME
DE
Artist
Issue date
COLOR FIGURE
Rev5
Dr. Jani
06/13/2013
1
Daniel MullerThe New England Journal of Medicine
permission.
reserved.
sounding board

cases from below 5% in 2000 to 45% in 2010,19
thereby reducing inappropriate antimalarial treat-
ment and improving community-based manage-
ment of fever and health outcomes.20 New point-
of-care tests also show promise. In Mozambique,
the use of such testing for CD4+ T cells at primary
health care clinics doubled the rate of initiation of
antiretroviral therapy and halved the time until
treatment initiation.21 Rapid, cost-effective NAT-
based testing for tuberculosis increased the rate
of case detection by up to 50% and reduced the
time until treatment initiation by a factor of
10.22,23 Other opportunities exist for point-of-care
testing to improve access to appropriate medical
services and thus patient outcomes. An accurate
test for preeclampsia may enable earlier diagno-
sis and appropriate care for a major cause of ma-
ternal death,24 and NAT or antigen testing for
HIV may improve the rate of pediatric case detec-
tion and treatment coverage, which is currently
below 50% in many low- and middle-income
countries.25
However, weak health systems limit the effect
of such testing programs. The initial adoption of
promising new diagnostics is hampered by slow
regulatory approval and uncertainty over how to
deploy new tests relative to existing technology.
This may lead to either inappropriate use or over-
use. After deployment, inventories of testing sup-
plies often run low, and the reliability of point-
of-care testing in real-life nonlaboratory settings
may be low.26 In addition, the use of such tests
has not always improved patient outcomes. For
example, rapid antenatal syphilis testing reduced

treatment delays but did not lead to higher treat-
ment rates or a reduction in perinatal mortality.27
The use of rapid tests for malaria has not always
improved the prescribing behavior of clinicians.28
Access to rapid tests for HIV did not improve the
uptake of same-day testing at antenatal clinics.29
Despite widespread use of rapid tests for HIV,
only 40% of HIV-infected persons know their sta-
tus, and 40% of those with positive test results do
not access follow-up care and may be at increased
risk for death or complications because of delayed
therapy.3
Study data suggest similar challenges with
new point-of-care technologies. Despite the avail-
ability of point-of-care testing for CD4+ T cells
at primary health care clinics in Mozambique,
only 30% of patients underwent same-day test-
ing, and 20% were not tested at all.21 Of those
tested and eligible for antiretroviral therapy, 40%
were lost to follow-up while undergoing addi-
tional testing and counseling before treatment.
The benefit of new point-of-care tests cannot be
taken for granted.
N e e d e d C h a n g e s in H e a lt h
S y s t e m s
Health systems have been designed around ei-
ther syndromic management or diagnostic test-
ing performed in the laboratory and are not well
adapted to the use of point-of-care testing. The
coming wave of such technologies demands
changes to health systems. We propose four key
areas where change is needed (Fig. 2).

First, testing policies need to be updated.
The World Health Organization (WHO) and
other normative bodies should provide recom-
mendations on how to use point-of-care tests
(including guidance on risks, benefits, and cost-
effectiveness), how to select the right products,
and where and how to deploy new technologies
in relation to existing tests. Even if such tests
are cost-effective, their use may incur additional
costs to health budgets, especially for new and
more sophisticated tests, and these implications
Figure 1 (facing page). Evolution of Point-of-Care (POC)
Diagnostic Testing.
Improvements in POC technology will lead to increas-
ingly complex tests run on devices that are smaller and
easier to use than the current generation of devices.
Panel A shows first-generation POC tests, which are
conducted with the use of simple chemical analyses
and devices. Most tests are lateral- or vertical-flow
devices that allow the specimen to flow across or
through the solid surface of the test strip past a reac-
tion area, resulting in a visual signal. Both manual and
automated readings of test results are common. Panel B
shows second-generation POC tests, which detect
more difficult diagnostic targets with the use of more
complex chemical analyses. Sophisticated, disposable
microfluidic test cartridges automate sample prepara-
tion and test processing. Cartridges are inserted into
small, portable instruments that automatically process
and read the results, which are displayed digitally. Pan-
el C shows the next generation of POC tests, which will
probably include more complex diagnostics for the si-
multaneous targeting of multiple diseases with the use
of instruments that are smaller and easier to use and

disposable devices that fully automate sample process-
ing, testing, and reporting of results. HIV denotes hu-
man immunodeficiency virus, and PCR polymerase
chain reaction.
permission.
reserved.
need to be made clear. For example, use of new
NAT-based testing for tuberculosis in South Af-
rica increased the cost of diagnosis by 55% and
the cost of treatment by 8%.30 Guidance on clini-
cal-management algorithms will also be needed.
Point-of-care tests will provide health care work-
ers, especially those in primary care and com-
munity-based settings, with unprecedented diag-
nostic information for guiding clinical decisions.
Governments, with guidance from the WHO, have
developed and implemented practical disease-
management algorithms that are based on clini-
cal judgment for primary health care settings in
which routine laboratory diagnostics were not
feasible or reliable. The increased availability of
accurate test data in these settings will necessi-
tate new clinical algorithms and guidance on how

to interpret and use diagnostic information. In
addition, point-of-care testing will increasingly
be used in the private health sector and in less
formal settings, such as pharmacies, retail outlets,
and homes. The increased availability raises con-
cern about product quality and testing perfor-
mance, and supportive but firm regulations on the
use of such tests in these settings will be needed.
Second, innovation will be needed in the de-
sign, operation, and workflow of clinics to ensure
that testing is accessible and results are used in
real time to guide treatment. Point-of-care test-
ing may lengthen clinic visits and place extra de-
mands on staffing and space. Bottlenecks at any
stage can increase waiting times and result in
extra visits by patients, and the benefits of on-
site testing may be lost. Clinics may need to hire
additional staff in key cadres, extend clinic hours
or work shifts, and change the scheduling of pa-
tients, clinic flow, and use of space in order to
facilitate onsite testing and immediate delivery
of follow-up care. Improved medical-record sys-
tems that capture test results and make them
available across different service departments may
improve the tracking and follow-up of patients.31
Steps should also be taken to increase testing
rates and reduce the effect of shortages in space
or test operators (e.g., use of multiplex or parallel
testing). The implementation of many of these
initiatives in public health systems will require
changes in government policy and resource al-
location — for example, to facilitate the exten-
sion of clinic operating times, hiring of addition-
al staff, and improvements in data-management

systems.
Third, systematic steps should be taken to
effectively decentralize point-of-care testing and
to improve the retention of patients both before
and after testing. Policies that enable new models
for expanded community-based testing and that
facilitate safe and reliable self-diagnosis provide
opportunities to better exploit such testing, as
well as drive technology innovation. Addressing
weaknesses in retention that persist despite on-
site testing will require initiatives both upstream
and downstream of the test to improve access to
testing and ensure appropriate linkage to follow-
up care. Interventions such as transportation and
food allowances for clinic visits and mobile-phone
reminders32 can help ensure that patients com-
plete their treatment as well as promote adherence
to clinical protocols among health care workers.33
Streamlining and integrating testing and related
services can improve access to treatment. For
Revised policy and normative guidance
Cost and cost-effectiveness of POC testing
Decentralization of services
Testing guidelines
Clinical algorithms
Community-based testing and self-testing
Improved operational
systems
Product regulation
Supply chain
Training
Quality assurance

Maintenance
Streamlined clinic services
New staff cadres and
shifts
Space reassignment
Patient scheduling
Improved medical records
Bundled procedures
Decentralization and
retention initiatives
Community-based testing
Self-testing
Linkage to care
Integrated services
Patient-centric services
Adherence tools
Figure 2. Health-System Improvements to Support
Expanded POC Testing.
Shown are four key areas of improvement in health
systems — revised policy and normative guidance,
improved operational systems, streamlined clinic ser-
vices, and decentralization and retention initiatives —
that will require strengthening in order to increase the
effect of POC testing. In these areas, the use of POC
testing may prompt system improvements that may
eventually extend beyond diagnostics to other areas of
health care.

permission.
reserved.
sounding board
example, the integration of outpatient services
and HIV therapy in Zambia resulted in increased
rates of case finding,31 and point-of-care testing
for CD4+ T cells in HIV screening clinics in
South Africa increased linkage to care.34 Other
opportunities exist across disease programs,
such as point-of-care tuberculosis testing in HIV
screening clinics or therapy centers.35 However,
the feasibility and cost-effectiveness of these in-
terventions need further investigation. Many
health-system innovations exist only in pilot
form. Translating useful pilots into policy and
routine practice is a challenge that requires in-
creased attention from both governments and
development partners.
Fourth, operational challenges to implement-
ing point-of-care testing need to be overcome.
Weaknesses in supportive services — including
product regulation, supply chain, human resourc-
es and training, quality assurance, and equipment
maintenance — are widespread and systemic.36
In particular, new in-service and preservice ini-
tiatives in training and retention of clinic staff

will be needed to ensure that new technologies
are used appropriately. Initiatives to improve these
areas are under way and can benefit other areas
of health care delivery.37 In addition, rational
planning for product uptake is necessary to en-
sure that the investment in point-of-care diag-
nostics is cost-effective and sustainable and that
tests are widely accessible.
D r i v in g t h e In n o v at i o n
in H e a lt h S y s t e m s
The rise of point-of-care testing is expected to
expand access to medical services, improve health
outcomes, and facilitate the sustainability of dis-
ease-control programs in low- and middle-income
countries. Although such technologies were ini-
tially focused on HIV, tuberculosis, and malaria,
they will be used in the diagnosis and treatment
of other diseases, and their deployment at scale
will require substantial investment. However, such
testing may not be cost-effective if the diagnos-
tic innovation is not matched with innovation in
health systems.
As point-of-care testing becomes more com-
mon in diagnostic medicine, it could drive this
innovation in health systems in at least three
ways. First, the supply of point-of-care tests will
directly induce changes, such as improved patient
flow within clinics. Second, the new technologies
tend to increase testing rates substantially, and
as more patients are tested, the demand for as-
sociated services will increase and existing sys-
temic weaknesses will be highlighted. This in-

creased pressure on health services will motivate
local and international initiatives to seek ways
to address such limitations. Finally, the enthusi-
asm for new point-of-care technologies among
public health practitioners, scientists, and the pri-
vate sector should elicit proactive efforts in re-
solving health-system bottlenecks so that the tests
can be successfully used.
There are many examples of system innovation
that have been prompted by the use of point-of-
care testing, such as the use of provider-initiated
HIV testing to increase diagnostic rates and im-
prove patient retention,38 wireless networks that
capture test data from remote sites and monitor
quality,39 and the “Test, Track, and Treat” pro-
gram for malaria, an international initiative of
the WHO designed to scale up malaria testing
linked to treatment and disease surveillance.19
However, more is needed to address the chal-
lenges described above. Commitment from gov-
ernments and global-health actors is necessary,
and strengthening of initiatives should be evi-
dence-based, drawing on operational research to
identify high-priority and cost-effective interven-
tions.40 The investment in developing new point-
of-care diagnostics has started to yield fruit. Now
health systems need to evolve to reap the benefits.
Disclosure forms provided by the authors are available with
the full text of this article at NEJM.org.
From the Instituto Nacional da Saúde, Maputo, Mozambique
(I.V.J.); and Clinton Health Access Initiative, Boston (T.F.P.).
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DOI: 10.1056/NEJMsb1214197

Copyright © 2013 Massachusetts Medical Society.
permission.
reserved.
Final Applied Lab Project (1 credit Lab Component)
Addresses course outcomes 1-5:
· apply the scientific method to scientific investigations
· state a scientific hypothesis and design a basic experiment
· conduct an experiment, make observations, and collect data
· use knowledge of biological principles to correctly interpret
qualitative and quantitative information
· use critical analysis to draw conclusions
This is the culminating assessment in BIOL 102. It is designed
to assess your ability to apply the principles of the scientific
method. For this project, you will complete the activity below.
Make sure to address all points (questions) associated with the
activity.
The Effect of low pH on Enzyme Activity
Design an experiment in which you will test the effect of an
acidic fluid on enzymatic activity. (Recall: enzymes are
proteins.) To complete this project, it may be useful for you to
review the Scientific Method Tutorial (found the Course
Content section of the classroom under the Science Learning
Center link) and the Scientific Method lab (Lab 1), so that you
can better understand how to design an experiment. It may also
be helpful for you to review your textbook and Lab 4
(Enzymes). As you review Lab 4, you will be reminded that

there are several factors that impact enzymatic activity: pH,
temperature, and amount of reagent. Feel free to refer to
observations and information from Lab 4 as you complete the
Final Applied Project (see the questions below). Or in other
words, it is OK to use the same enzyme/subtrate/method as you
did in lab 4 (but modify the treatment), or you can search on-
line to find a different enzyme/subtrate/method for measuring
enzyme activity.
As you design your experiment for this project, please
remember that you are trying to examine how an acidic fluid
will modify the outcome of an enzymatic reaction. To
successfully complete this project, you will need to identify the
question(s) being asked in your experiment and the hypothesis
that you are testing. In your experimental design, you must
clearly explain what you are doing. That means that you will
need to identify the enzyme and the acid, as well as explain
your experimental protocol (this information will help you to
answer question 2). You must also thoroughly explain how the
addition of the acidic fluid impacted the overall reaction
process (this information will help you to answer question 4).
Hint: Keep in mind that the acid will change the environmental
conditions of the experiment (for example, a low pH value
could change the shape of the active site on the enzyme
protein), without directly participating in the reaction.
Lab Materials
You may need all or some of the following, depending on your
experimental design:
Materials from your lab kit:
· pH paper
· hydrogen peroxide solution (you can purchase this at a
pharmacy if you have used up the bottle that came with the lab
kit)
· plastic beakers or cups
· vinegar
· yeast (can be purchased at grocery store if you need more)
· balloons

· plastic bottle
· marker for labeling of beakers
You may choose to use additional materials (different acidic
solutions and/or different organisms and/or differnet subtrate(s)
if you chose to look at an enzyme other than catalase).
Outline submit in assignment folder in week 7.
I suggest you include the following in your outline:
· Name of enzyme you will use, and source (organism)
· The substrate
· How you will measure enzyme activity (method)
· What type of treatment you will you; type of solution(s), pH,
length of exposure, how you will treat your samples
· The control(s) in the experiment
· Sample size
· Maybe how you plan to present your data (table and/or type of
graph)
· Anything else you would like to get feedback on before you
start your experiment.
Write a paper that includes the following:
1. Title page: title, your name, course name, semester
2. Introduction: introduce your project, include needed
background information, the question(s) that you are asking and
a clear hypothesis for your experiment.
3. Design an experiment. Provide a detailed account of the
materials and methods used to conduct the experiment. Also
include the methods for data collection and analysis.
4. Conduct the experiment and record your results. What did
you observe? Present your data in table and/or graph .
Remember to include the unit of measure.
5. Use your knowledge about enzymes and acids to interpret and
discuss your results. It may be necessary for you to refer to
your textbook and/or use other information resources. What
effect does the acidic treatement have on the enzyme activity?
Looking back, how could you have improved your experiment?
6. What is your conclusion? Was your hypothesis supported?
7. Cite all reference sources used (including text book) and

provide a reference section with citations in APA format
Submission
Submit your final applied lab project as an attached Word
document in the assignment folder by the due date specified in
the course schedule
THE LANCET
1886 Vol 349 • June 28, 1997
(seven partial or complete responses, four stable disease for
more than 6 months), while 12 (52%) did not. The median
time to progression for responding patients was 11 months
(range 5–29) and for non-responding patients 3 months
(range 0–6). The rate of response did not depend on the type
of chemotherapy given. There was no difference between
responders and non-responders in age, lymph node status, or
recurrence-free period. Two of eight (25%) postmenopausal
patients responded, compared with nine of 15 (60%)
premenopausal patients. The incidence of MRP-positive
tumours was not different for patients with soft tissue (one
of three), bone (two of eight), or visceral metastases (five of
twelve) as predominant site of relapse. In patients receiving
first-line chemotherapy, MRP was more often positive in
non-responding tumours (50%) than in responding tumours
(18%). Only one of eight (13%) MRP-positive tumours had
an objective response (5 months), compared with six of 15
(40%) MRP-negative tumours. Analysing for overall
response, including stable disease, two of eight (25%) MRP-
positive tumours responded, compared with nine of 15
(60%) MRP-negative tumours (odds ratio: 0·22; 95% CI
0·03–1·49). Patients with MRP-positive tumours showed a
shorter time to progression on first-line chemotherapy than

those with MRP-negative tumours (Cox proportional hazard
model, p=0·006, figure). The relative hazard rate for time to
progression in patients with MRP-positive tumours,
compared with MRP-negative tumours, was 4·08 (95% CI
1·50–11·12). At 9 months, all eight patients with MRP-
positive tumours showed disease progression, while seven of
15 of those with MRP-negative tumours did not (four
objective responses, three stable disease). In Cox
multivariate analysis for time to progression, MRP was the
only significant variable in the model.
Of the 41 patients who received chemotherapy after one
or more lines of hormonal therapy, 19 (46%) responded
(seven partial responses, 12 stable disease), whereas 22
(54%) did not. In these patients, there was no significant
difference in the rate or duration of response, or in the time
to progression between patients with MRP-positive and
MRP-negative tumours, suggesting differences in tumour
cell biology. Metastatic breast cancer patients who receive
chemotherapy as the first choice of treatment usually are
premenopausal, are oestrogen-receptor and progesterone-
receptor negative, and may have visceral metastases. These
are all unfavourable prognostic factors. Women first treated
with hormonal therapy are usually postmenopausal, have
receptor positive tumours, and have bone rather than visceral
metastases. We conclude that MRP expression is an
important predictor of poor prognosis in patients with breast
cancer who were treated with chemotherapy as first-line
systemic therapy for recurrence.
We thank Maxime Look for statistical analysis. This work was
supported
by the Dutch Cancer Society (Grants DDHK95-1051, DDHK96-
1236).

1 Cole SPC, Bhardwaj G, Gerlach JH, et al. Overexpression of a
transporter gene in a multidrug-resistant human lung cancer cell
line.
Science 1992; 258: 1650–54.
2 Flens MJ, Zaman GJR, van der Valk P, et al. Tissue
distribution of the
multidrug resistance protein. Am J Pathol 1996; 148: 1237–47.
3 Nooter K, Westerman AM, Flens MJ, et al. Expression of the
multidrug resistance-associated protein (MRP) gene in human
cancers.
Clin Cancer Res 1995; 1: 1301–10.
Departments of Medical Oncology (K Nooter) and Clinical
Pathology and
Division of Endocrine Oncology, University Hospital Rotterdam
and
Rotterdam Cancer Institute, PO Box 2040, Rotterdam,
Netherlands
100
80
60
40
20

0P
ro
g
re
ss
io
n
-f
re
e
s
u
rv
iv
a
l
(%
)
0 3 6 9 12 15 18 21 24 27 30
15 12 7 6 5 2 1 1 1 1 0
Time (months)
p=0·006
RHR (95% CI) 4·08 (1·50–11·12)

MRP-
positive (n=8)
MRP-negative (n=15)
MRP-negative
8 3 1 0 0 0 0 0 0 0 0MRP-positive
Time to progression for patients treated with first-line
chemotherapy for recurrence as a function of MRP status.
Patients at risk at start and at every 3 months are indicated
RHE=relative hazard rate.
Complement C3 and factor B
cerebrospinal fluid concentrations in
bacterial and aseptic meningitis
Philip F Stahel, David Nadal, Hans-Walter Pfister,
P Maria Paradisis, Scott R Barnum
Establishing a diagnosis is difficult in most cases of acute
meningitis, since its clinical signs are non-specific, and
laboratory examination of cerebrospinal fluid (CSF) often
does not accurately differentiate between bacterial and
aseptic meningitis.1,2 Therefore the identification of a
discriminating parameter, which might lead to a rapid and
accurate clinical test, would be of value in the differential
diagnosis of acute meningitis.
Several studies have suggested that the complement system
contributes to intrathecal inflammation in bacterial
meningitis.3 In a retrospective analysis, we measured the
concentrations of the alternative pathway complement
components C3 and factor B in CSF samples obtained by
lumbar puncture from 39 patients with clinically suspected

acute infectious meningitis, and from 64 controls without
central nervous system infection, with an ELISA developed in
our laboratory. 18 patients (median age 40 years; range
14–67 years; 9 female) were diagnosed as having bacterial
meningitis, on the basis of positive bacterial culture or on
detection of bacterial antigen in CSF. The pathogens were
Streptococcus pneumoniae (n=10), Haemophilus influenzae (3),
Neisseria meningitidis (3), Listeria monocytogenes (1), and
Streptococcus bovis (1). 21 patients were diagnosed as having
aseptic meningitis (median age 8 years; range 2 months to 13
years; 6 female) on the basis of CSF pleocytosis with a
predominance of mononuclear cells, negative bacterial and
fungal CSF and blood cultures, negative results on CSF
antigen detection tests for S pneumoniae, H influenzae, and
N meningitidis, and full recovery without antibiotic treatment.
No patient had received antibiotics or steroids before
diagnostic lumbar puncture, and all CSF samples were
obtained on admission. The mean C3 concentration in the
CSF of patients with bacterial meningitis (48·32 [SD 50·74]
µg/mL) was significantly higher than in aseptic meningitis
(2·16 [1·82] µg/mL; p<0·001, Wilcoxon rank sum test) or in
controls (2·49 [2·18] µg/mL; p<0·001). Similarly, factor B
CSF concentrations were significantly raised in patients with
bacterial meningitis (15·89 [17·36] µg/mL) compared with
those with aseptic meningitis (0·25 [0·20] µg/mL; p<0·001)
or controls (0·29 [0·26] µg/mL; p<0·001). C3 and factor B
CSF concentrations in bacterial meningitis did not correlate
THE LANCET
Vol 349 • June 28, 1997 1887
Nonsense mutation of prostacyclin
synthase gene in a family

Tomohiro Nakayama, Masayoshi Soma, Dolkun Rahmutula,
Yoichi Izumi, Katsuo Kanmatsuse
We found a nonsense mutation in exon 2 of the human
prostacyclin-synthase gene in a family with essential
hypertension and cerebral infarction. Prostacyclin (PGI2) is
an inhibitor of platelet aggregation, smooth muscle cell
proliferation, and vasoconstriction. Prostacyclin synthase
(PGIS), which catalyses the formation of PGI2 from
prostaglandin H2, is widely distributed, predominantly in
vascular endothelial and smooth muscle cells. We have
reported the organisation of this gene.1
We searched for possible point mutations in the exons
using peripheral blood from 100 patients with essential
hypertension by PCR and single strand conformation
polymorphism (PCR-SSCP) analysis. One patient had an
abnormally migrating band on exon 2. Sequencing of this
exon showed a nonsense mutation in codon 26
(CGA/TGA). This nucleotide change makes Bst EII the
restriction site. 300 people (150 with essential hypertension
and 150 healthy controls) were screened by PCR and Bst EII
digestion. The mutation was found in one patient with
essential hypertension and in none of the controls. The
patient was shown to be heterozygous for this mutation. This
mutation of the stop codon is 76 bp downstream from ATG,
the start codon in cDNA, thus a large part of mRNA,
Male
Female
Dead
HT

CI
Hypertension
Cerebral infarction
Patient first detected
66y
HT
69y
HT
66y
HT
61y
HT
68y
Apoplexy
HT
63y
HT
CI
57y

HT
CI
–
The 247-bp fragment digested with Bst EII will give
fragments of 189 bps, 58bps
– + +– + +
247bp
189bp
58bp
Mutation
Family tree and PCR
with CSF total white blood cell counts or CSF protein
concentrations (r<0·6, Spearman’s rank correlation
coefficient).
We found that complement concentrations in the CSF may
be of clinical value in distinguishing bacterial from aseptic
meningitis. With cut-off levels of the mean value +2 SD for
the aseptic meningitis population (5·8 µg/mL for C3 and 0·65
µg/mL for factor B; figure), C3 and factor B CSF
concentrations were highly sensitive (both 100%) and highly
specific (95·2% and 100%, respectively) tests for the
diagnosis of bacterial meningitis, and associated with a
negative predictive value of 100%, and a positive predictive
value of 94·7% (C3) and 100% (factor B). Quantification of

C3 and factor B CSF concentrations can be completed
within 3–4 hours. We plan to test these preliminary results in
a multicentre prospective study.
1 Lindquist L, Linné T, Hansson L-O, Kalin M, Axelsson G.
Value of
cerebrospinal fluid analysis in the differential diagnosis of
meningitis:
1000
100
10
1
0·1
0·01
1000
100
10
1

0·1
0·01
C
3
(
µg
/
m
L)
F
a
c
to
r
B
(
µg
/
m
L)
Bacterial
meningitis

Aseptic
meningitis
Controls
Mean value
Cut-off
Complement C3 and factor B concentrations in the CSF of
patients with infectious meningitis and controls
Each point represents the mean of duplicate sample analysis.
Cut-off
level=mean of aseptic meningitis group+2 SD for differentiation
between bacterial and aseptic meningitis.
a study in 710 patients with suspected central nervous system
infection.
Eur J Clin Microbiol Infect Dis 1988; 7: 374–80.
2 Rodewald LE, Woodin KA, Szilagyi PG, Arvan DA, Raubertas
RF,
Powell KR. Relevance of common tests of cerebrospinal fluid in
screening for bacterial meningitis. J Pediatr 1991; 119: 363–69.
3 Stahel PF, Barnum SR. Bacterial meningitis: complement gene
expression in the central nervous system.
Immunopharmacology. (In
press)
Department of Microbiology, University of Alabama at
Birmingham,
Birmingham, AL 35294, USA (S Barnum); Unit of Infectious
Diseases,

University Children’s Hospital, Zürich, Switzerland; and
Department of
Neurology, Klinikum Grosshadern,University of Munich,
Munich, Germany
THE LANCET
1888 Vol 349 • June 28, 1997
Colonic perforation and serosal
tears associated with colonoscopy
Yoshiharu Uno, Takayuki Morita
Serosal tears with no mucosal damage are a complication of
colonoscopy.1–4 These tears have been attributed to the
pressure of the air introduced through the colonoscope or to
stretching the wall of the colon.
A colonoscope (CF230I, Olympus Inc, Japan) was pressed
against the mucosa of a piece of sigmoid colon removed
during surgery for rectal cancer at a continuous pressure of
2–3 kg/cm2 (figure, a). First, the muscularis propria ruptured
(figure, b), and then the serosa tore (figure, c), before the
mucosa ruptured and the colonoscope penetrated the wall
(figure, d). We repeated this experiment at 40 different
points (ten points in each of four segments: sigmoid,
descending, transverse, and ascending colon) of resected
colons from eight patients with mean age 62 (SD 11) years.
Perforation occurred in the above order, regardless of age,
colonic segment, or sex.
We asked 60 doctors in our hospital how injury occurred
when the tip of a colonoscope was pressed against the
colonic wall. They all replied that perforation started from

the mucosa, proceeded to the muscularis propria and then
the serosa. Our results suggest that, when the tip or a bend
of a colonoscope is pressed hard against the colonic mucosa,
a seromuscular tear will probably occur, even when there is
no mucosal injury. If the mucosa does not rupture,
peritonitis is unlikely.
1 Livstone EM, Cohen GM, Troncale FJ, Touloukian RJ.
Diastatic
serosal lacerations. Gastroenterology 1974; 67: 1245–47.
2 Livstone EM, Kerstein MD. Serosal tears following
colonoscopy.
Arch Surg 1976; 111: 88.
3 Kozarek RA, Earnest DL, Silverstein ME, Smith RG. Air-
pressure-
induced colon injury during diagnostic colonoscopy.
Gastroenterology
1980; 78: 7–14.
4 Ehrlich CP, Hall FM, Joff N. Postendoscopic perforation of
normal
colon in an area remote from instrumentation. Gastrointest
Endosc
1984; 30: 190–91.
First Department of Internal Medicine and Second Department
of
Surgery, Hirosaki University School of Medicine, Aomori 036,
Japan
(Y Uno)
Perforation of colon by colonoscope
a, b, c, d depict how serosal tears occur as complication of
colonoscopy.

6 kbp,2 can not be translated. Consequently the activity of
PGIS may be decreased.
The patient is a 57-year-old woman with essential
hypertension who presented with a blood pressure of
177/113 mm Hg. Her electrocardiogram showed left
ventricular hypertrophy. Although she had never smoked and
rarely consumed alcohol, at the age of 50 she had a transient
ischaemic attack. We looked for this mutation in her family.
Her father died of stroke aged 70. Her mother was healthy
until her death at age of 92. She had eight siblings including
two elder brothers one of whom died in the war and the
other of chronic renal failure. Three of the five living siblings
had the mutation (figure); all were hypertensive. One with
the mutation had had a cerebral infarction. As essential
hypertension is thought to be a multifactorial disorder, PGIS
may be one of the genes involved.
1 Nakayama T, Soma M, Izumi Y, Kanmatsuse K. Organization
of the
human prostacyclin synthase gene. Biochem Biophys Res
Commun 1996;
221: 803–06.
2 Miyata A, Hara S,Yokoyama C, Inoue H, Ullrich V, Tanabe T.
Molecular cloning and expression of human prostacyclin
synthase.
Biochem Biophys Res Commun 1994; 200: 1728–34.
Second Department of internal Medicine, Nihon University
School of
Medicine, Ooyaguchikamimachi 30–1, Tokyo 173, Japan (T
Nakayama)

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