CareEvolveOption 1 CareEvolve portal: NEW!! Updated CareEvolve Instructions
Basic functionality included
This option is already included in the price.
CoVerifiedOption 2 CoVerified campus management platform NEW!! Onboarding Website
|Detailed information around upcoming webinars, sign up process, timelines, and security documentation can be found at www.coverified.us/broad.|
Ready-to-ship tube with properly placed swab and barcode (from CRSP-provided label).
Test turnaround time: within 24 hours of receipt at Broad
Students can login and register for results at: https://docs.google.com/presentation/d/1izY-518Esba1tcTcwtR05SZNvNrURvqDhXN-x86Qedk/edit#slide=id.g8bd924a234_2_0
If you have any questions regarding your test results, please contact your COVID test site administrator.
Regardless of option chosen above, samples may be rejected or significantly delayed if they arrive with any of the following nonconformances:
*updated July 28, 2020
Unsatisfactory reason codes that only apply to samples submitted in media are NOT italicized.
Possible Unsatisfactory/“Test Not Performed (TNP)” Reason Codes for samples submitted using dry, anterior nasal swab collection (aka NO media) OR in media are italicized.
TNP reasons found in CareEvolve are shown in bold/italics.
When the laboratory issues an unsatisfactory or TNP result in a report, the individual must be tested again and a NEW sample must be submitted to the laboratory in order to receive a confirmed test result.
|Result||Definition ("reason" or column J in the results file)||Description|
|NEGATIVE||2019-novel Coronavirus (2019-nCoV) not detected by the qRT-PCR assay. Consider testing for other respiratory viruses or re-collecting for 2019-nCoV testing. Note: Optimum timing for peak viral levels during infections caused by 2019-nCoV have not been determined. Collection of multiple specimens from the same patient may be necessary to detect the virus.||Applies to both assays with and without media.|
|POSITIVE||Positive for detection of 2019-novel Coronavirus (2019-nCoV) by qRT-PCR.||Applies to both assays with and without media.|
|INVALID||This specimen failed to produce a valid result. An invalid result means no nucleic acids (viral or human) were detected by qRT-PCR. Consider re-collection of specimen.||Applies to both assays with and without media.|
|CANCELED||Testing canceled per submitter.||Canceled by ordering provider.|
|UNSATISFACTORY_1||Unsatisfactory for 2019-novel Coronavirus (2019-nCoV) testing by PCR: improper specimen transport medium.||Applies to samples submitted in media. The media cannot be pipetted (too viscous).|
|UNSATISFACTORY_2||Unsatisfactory for 2019-novel Coronavirus (2019-nCoV) testing by PCR: inappropriate timing of collection relative to specimen receipt. Specimens must be received within 72 hrs of collection unless frozen.||Samples are received more than 72 hours after collection date|
|UNSATISFACTORY_3||Unsatisfactory for 2019-novel Coronavirus (2019-nCoV) testing by PCR: sample tube unlabeled.||Sample tube is unlabeled and cannot be matched to an order.|
|UNSATISFACTORY_4||Unsatisfactory for 2019-novel Coronavirus (2019-nCoV) testing by PCR: insufficient volume.||Applies to samples submitted in media. The tube is either empty or does not contain enough media volume to perform the test.|
|UNSATISFACTORY_5||Unsatisfactory for 2019-novel Coronavirus (2019-nCoV) testing by PCR: tube sample data does not match the submission form.||This is used when the order and tube label do not match.|
|UNSATISFACTORY_6||Unsatisfactory for 2019-novel Coronavirus (2019-nCoV) testing by PCR: laboratory accident.||This is used when a test can not be performed due to a laboratory error and the sample must be resubmitted.|
|UNSATISFACTORY_7||Unsatisfactory for 2019-novel Coronavirus (2019-nCoV) testing by PCR: leaking/broken tube.||This is used for broken or leaking tubes. Very similar to reason #10, but this is used for samples submitted in media.|
|UNSATISFACTORY_8||Unsatisfactory for 2019-novel Coronavirus (2019-nCoV) testing by PCR: sample not received.||This is used when we have an electronic or physical order in hand but do not receive a physical specimen.|
|UNSATISFACTORY_9||Unsatisfactory for 2019-novel Coronavirus (2019-nCoV) testing by PCR: incomplete requisition.||This is used when required elements of the requisition are missing and we are unable to obtain the information before the sample expires, or if there is not enough information to know who to reach out to for the missing information.|
|UNSATISFACTORY_10||Unsatisfactory for 2019-novel Coronavirus (2019-nCoV) testing by PCR: Specimen tube received uncapped or broken.||This is used in cases where the sample arrives in an uncapped or broken tube.|
|UNSATISFACTORY_11||Unsatisfactory for 2019-novel Coronavirus (2019-nCoV) testing by PCR: Sample unsuitable for automated processing.||The first step in processing is to add media to the swab. After adding media, some samples are too viscous to process. This most often happens when the patient does not blow their nose before collecting the sample, resulting in excessive mucus. This reason may also be used if we cannot read the barcode on the tube.|
|UNSATISFACTORY_12||Unsatisfactory for 2019-novel Coronavirus (2019-nCoV) testing by PCR: Swab collection not to submission standards.||This is used when the swab is upside down in the tube, there are multiple swabs in the tube, or when there is no swab in the tube.|
The Broad CRSP SARS-CoV-2 RT-PCR Diagnostic Assay is used for in vitro qualitative detection of SARS-CoV-2 in respiratory specimens.
This means the test is designed to detect the presence of the genomic material of the SARS-CoV2 novel coronavirus, the pathogen responsible for the disease known as COVID-19.
The test is performed at the Clinical Research Sequencing Platform (CRSP) which is the clinical testing laboratory at the Broad Institute of MIT and Harvard. This laboratory operates under a set of government regulations known as CLIA that allow us to run high complexity molecular tests, such as the one used to detect coronavirus. The test that is run by CRSP is a modified version of a test developed by the CDC. Our laboratory has tested approximately 250,000 people for the presence of coronavirus so far.
The test we have developed for the Safe For School program incorporates many improvements to the process that we have made in response to our experience and observations in running a quarter of a million tests so far. In place of the narrow swabs that are placed deeply into the upper nasal cavity (nasopharyngeal swabs) we now use a swab that is much shorter and can collect sample from the lower nasal cavity (anterior nares swabs). This is a much more pleasant experience for the person being tested. The swab is also placed into a tube without liquid (dry swabbing) as this allows the laboratory more flexibility in getting the most material off the swab for testing as possible. Our laboratory has performed validation studies to confirm that this swab type and collection method perform similarly (and in fact are better) than the wet swab collection method.
After the swab arrives at our laboratory, the collected material (human cells and virus particles, if present) are reconstituted off the swab using our automated liquid handling robots. The material is treated to inactivate any virus present (making it safer) and the cells are broken up to release the genetic material within. The genetic material is then amplified with probes that target either the virus genome or a human gene. We test for the presence of a human gene to confirm that the swab was successfully taken and that the testing process is working. If viral particles are present they are amplified in a way that produces a signal that can be detected on a specialized piece of equipment known as a real-time PCR machine. The data from this instrument is sent to our pipeline which interprets the signal as the presence or absence of the virus and checks the validity of the test. A test report is then created and returned.
The limit of detection (LoD) is defined as the lowest concentration at which 19/20 replicates (or approximately 95% of all true positive replicates) are positively detected. Initial LoD estimates were made based on a dilution series of a synthetic SAR-CoV2 construct brought all the way through the process. The preliminary LoD established by the dilution series was confirmed by testing a total of 20 replicates of the Synthetic SARS-CoV-2 RNA Control at the varying copy levels (Table 1).
|Concentration||# positive replicates/total # replicates||% True Positives Detected|
|100 copies (2.67 copies/µL)||19/20||95%|
|60 copies (1.6 copies/µL)||19/20||95%|
|40 copies (1.1 copies/µL)||18/20||90%|
|20 copies (0.5 copies/µL)||14/20||70%|
|10 copies (0.3 copies/µL)||13/20||65%|
|5 copies (0.15 copies/µL)||4/20||20%|
The LoD for this assay was determined to be 60 viral copies into RNA extraction (1.6 copies/µL). 19/20 extraction replicates of RNA construct spiked into human cell controls tested at this concentration (19/20) were positive.
30 positive specimens and 28 negative specimens, from upper respiratory swabs with positive SARS-CoV-2 results or negative test results previously tested using the CRSP v1 assay. Additionally, 20 replicates at ~2X of the LoD were also run through the process from extraction to detection. In order to be considered passing, there had to be 95% agreement at 1x-2x LoD, and 100% agreement at all other concentrations and for negative specimens (Table 2). Both evaluations passed.
|Samples||Mean Ct (SD)||% Agreement (# Pos or Neg)/Total|
|SARS-CoV-2 Positive Clinical Samples||20.48 (SD:4.43)||100% (30/30)|
|SARS-CoV-2 Positive Clinical Samples at 2X LoD||32.39 (SD: 0.87)||95% (19/20)|
|Negative Clinical Samples||N/A||100% (28/28)|
Spun polyester swabs were coated in a negative clinical matrix and allowed to dry. A high viral load positive clinical specimen was diluted to create a range of copies/mL at between 1-3X the LoD of the assay and spiked on to each swab. Swabs were processed through the dry swab reconstitution, extraction, and detection protocols. LoD was determined as the level at which 19/20 replicates were successfully called positive (Table 3). The LoD was confirmed at 60 copies/µL.
|Concentration||# positive replicates/# total replicates||Avg N Ct||Std Dev Ct|
The Broad CRSP v2 assay (high throughput) with dry swabs as an input is validated with an LoD of 1.6 copies/µL (or 1600 copies/mL).
For context, a study published in The Lancet1 found median viral load in COVID positive patients at presentation to be 158,489 copies/mL.
The ability to detect the virus is impacted not just by the performance of the test, but also by the timecourse of the infection in an individual.
Researchers have shown2 that the longer in time after the onset of symptoms a person is tested, the lower the chance of detecting the virus in that person (as they have largely cleared the virus out of their system).
The absolute sensitivity and specificity of the type of testing we and others are doing for COVID-19 diagnostics are difficult to calculate, as there is no gold standard assay against which we can compare results.
In the absence of this, the FDA and clinical best practice guidance is to compare the test on clinical specimens with positive and negative results that have been established using another FDA-approved test.
To this end, we initially validated our assay with samples that had been run at another laboratory using their FDA-approved test.
This validation cohort included 103 positives and 40 negative samples.
Further, 20 of the positive samples were diluted to the established limit of detection of the assay (the lowest concentration of virus at which we see 95% accuracy in test results).
In all cases, we saw 100% concordance with the established result.
Subsequently, we ran 543 cases through our assay and a collaborating clinical labs FDA-approved assay and again saw 100% concordance of results.
In additional studies where we have run a range of clinical samples several times internally, we have seen sensitivity and specificity >95% when we are at or above our limit of detection.
Studies have now shown that the infectivity of an individual relates to their viral load and that the viral load and infectivity are both highest in the few days immediately preceding and following the onset of symptoms.
When we see a low viral load we believe it is most often a person who has been infected but has largely cleared the virus and may therefore be less infectious.
When there are large numbers of positive individuals in a tested population, the main mode of test failure that we are concerned with is False Negatives (a negative test result for a person who does in fact have the virus). When the tested population has a low viral prevalence then we become more concerned with False Positives (a positive result in a person who does not have the virus). Since our validation cohorts did not show evidence of any false positives we have a high degree of confidence that false positives are not a significant source of error in our assay. Based on data from recent weeks in large cohorts where the prevalence is expected to be very low, we validate this observance. In one institution with >5000 tests run between mid-May and mid-June, we find only 3 positives. In each of the individuals who tested positive we have reason to believe that they were in fact exposed to the viral material being tested for (therefore in testing terms, we consider them True Positives). This empirical data would suggest that the assay False Positive Rate can be no higher than 1/5000 (or 0.02%) and may in fact be lower. Given the significant implications of False Positives, especially within a campus setting, we plan to routinely repeat testing of some proportion of positive results to further ensure the very high specificity of our test.