Post-operative Changes in Children With Severe Neurological Impairment
Conditions: Post-operative Functional Decline, Severe Neurological Impairment
Sponsor: University of British Columbia

Protocol 
  
Post-Op Changes Study  
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Post-operative Functional Changes in Children  
with Severe Neurological Impairment  
 
Study Protocol 
 
 
Principal Investigators  
Dr. Hal Siden 
 
 
 
 
Department of Pediatrics, UBC 
 
 
 
 
BC Children’s Hospital Research Institute 
 
 
 
 
Phone: 604 875 2776 
E-mail: hsiden@bcchr.ca  
 
 
 
 
 
Dr. Liisa Holsti 
Department of Occupational Science and Occupational Therapy, UBC 
Department of Pediatrics, UBC 
BC Children’s Hospital Research Institute 
 
 
 
 
Phone: 604 822 2000 ext. 5200 
E-mail: liisa.holsti@ubc.ca  
 
Co-Investigators 
 
Dr. Kishore Mulpuri 
Department of Orthopaedics, UBC 
Pediatric Orthopaedic Surgery, BCCH 
Phone: 604 875 2054 
E-mail: kmulpuri@cw.bc.ca 
 
 
 
 
 
Maria Juricic 
Department of Physical Therapy, UBC 
Physiotherapy, BCCH 
Phone: 604 875 2345 ext 7274 
Email: mjuricic@cw.bc.ca 
 
 
 
 
 
Stacey Miller 
Department of Physical Therapy, UBC 
Child Health BC Hip Surveillance Program for Children with Cerebral 
Palsy 
Phone: 604 875 2345 ext 4099 
Email: smiller4@cw.bc.ca 
 
 
 
 
 
Daphne O’Young 
Department of Orthopaedics, UBC 
Occupational Therapy, BCCH 
Phone: 604 875 2345 ext. 5443 
Email: doyoung@cw.bc.ca 
 
Dr. Sophie Stukas 
Department of Pathology and Laboratory Medicine, UBC 

Protocol 
  
Post-Op Changes Study  
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BC Children’s Hospital Research Institute 
Email:sophie.stukas@ubc.ca   
 
 
Collaborators  
 
Dr. Ramon Klein Geltink 
Department of Pathology and Laboratory Medicine, UBC 
BC Children’s Hospital Research Institute 
Email: ramon.kleingeltink@bcchr.ca  
 
Dr. Cheryl Wellington 
Department of Pathology and Laboratory Medicine, UBC 
BC Children’s Hospital Research Institute 
Email: cheryl.wellington@ubc.ca   
 
Dr. Andrea Simmonds 
Department of Orthopaedics, UBC 
Pediatric Orthopaedic Surgery, BCCH 
Email: asimmonds@cw.bc.ca 
 
Research Assistant 
Elaha Niazi 
 
 
 
 
BC Children’s Hospital Research Institute 
 
 
 
 
Email: elaha.niazi@bcchr.ca  
 
Contact 
 
 
Anne-Mette Hermansen 
BC Children’s Hospital Research Institute 
Phone: 604 875 2000 ext. 6909 
E-mail: ahermansen@bcchr.ca 

 
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BACKGROUND 
 
Children with Severe Neurological Impairment (SNI) have significant neurological disorders, either genetic 
or acquired in origin1, that result in developmental impairments influencing intellectual function, muscle 
tone, mobility and communication2. These children have ongoing health complexities that often require 
surgical procedures to manage their underlying condition, reduce pain burden, and improve functional 
limitations3,4. Examples of surgeries include spine stabilization, tendon releases, and gastrostomy.  
 
Children undergoing major surgical interventions are at risk of developing acute onset and persistent 
postoperative functional decline (PoFD)5, regardless of their pre-existing cognitive state. While evidence 
for PoFD exists, it has predominantly been reported in the frail elderly or focused on children without 
SNI6–11. To explore this phenomenon more systematically in children with SNI, we conducted qualitative 
interviews with 12 caregivers to investigate prolonged PoFD. Based on retrospective recall, 80% of 
caregivers reported unexpected, long-term PoFD in their children13. The changes occurred across a wide 
range of indicators; no two children were alike, making the use of broader categories of functional  
indicators of change challenging. Two previous studies examining the impact of hip and spinal surgery 
on health-related quality of life (including functional outcomes)14,15 found a decrease in quality of life at 
six weeks post-surgery, followed by gradual improvement at 12 months and significant improvement 
five years after surgery14,15. However, these studies analyzed broader categories of function, without 
specific item-level quantification during the perioperative period. Furthermore, no prospective studies 
have systematically and specifically examined PoFD in a larger cohort of children with SNI. 
 
While there are several factors hypothesized to contribute to PoFD, including prolonged anaesthesia 
exposure16–19 and ongoing pain20, emerging evidence suggests that inflammation due to surgery may be a 
more likely mediator. Surgery triggers a local inflammatory response with the potential to reach the 
brain21–23. Meta-analyses have found that certain inflammatory markers (eg. C-reactive peptide [CRP], 
interleukin-6) are significantly elevated in elderly patients with PoFD vs those without1,2, and in adult mice, 
inhibiting inflammatory cytokines attenuates PoFD-related effects on cognition3,4. Children with SNI may 
be especially susceptible to inflammation-induced PoFD due to their neurological vulnerability and 
potentially dysregulated immune response5. Inflammation due to surgery may thus be a factor which 
interacts with neuronal processes to possibly contribute to PoFD. However, the perioperative 
inflammatory profile in children with SNI and its potential connection to PoFD has not yet been 
systematically studied. 
 
Given the documented experiences with medical trauma in this group6, we must identify family-informed 
methods before conducting large-scale work involving non-medically necessary data collection. 
Therefore, the purpose of this pilot study is to test the feasibility and acceptability of conducting an 
observational to explore PoFD in children with SNI.  The ability to identify the best methods for evaluating 
potential functional decline (temporary or persistent) in children with SNI will assist in finalizing a study 
protocol for a planned larger study. 
 
SIGNIFICANCE AND JUSTIFICATION 
 
To our knowledge, this will be the first study to prospectively and quantitatively document item-level 
post-operative changes in function and quality of life for children with SNI and their caregivers. Expanding 
upon previous research14,15 that has obtained a total score and sub-domain scores of function/health-
related quality of life, this work aims to examine more specific, item-level details regarding function pre- 

 
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and post-operatively. Additionally, no prior research has investigated potential mechanisms of PoFD, 
including inflammation, in children SNI.  
 
This pilot study will help refine methods for a planned larger study on this phenomenon. The 
implications of this future work may significantly improve peri-operative care for children with SNI. Pre-
operatively, this could assist clinicians in guiding families through surgical decision-making7 and the post-
operative recovery trajectory. Post-operatively, our future findings could inform the development of 
targeted, clinically appropriate, postoperative rehabilitation goals as well as help identify needs for 
multidisciplinary health and community supports. This work will also set a foundation for research that 
investigates potential mechanisms for the changes in functional status, if present, which could inform 
interventions that ameliorate or prevent PoFD. Given the incurable nature of disorders that cause SNI, 
the impact of SNI and its functional implications on both the child and family is profound26–28. This line of 
work thus has implications to ultimately enhance the quality of life in children with SNI and their 
families. 
 
STUDY OBJECTIVES 
 
In this pilot study, we will evaluate the feasibility and acceptability of our protocol to explore the 
functional post-operative trajectory in a developmentally and neurologically impaired pediatric 
population. We will evaluate consent rate, protocol delivery and outcome completion, and acceptability. 
We will also describe common elements of the experience of these children and their families, including 
information from a functional skills questionnaire, parental descriptions of pre-post abilities, and 
parental descriptions of their own quality of life. The objectives of our research are: 
 
Primary outcomes 
 
Objective 1: To evaluate the feasibility and acceptability of our study protocol for children and families 
living with SNI.  
 
To evaluate feasibility we will use the following evaluation points: 
1. Consent rate: The overall average consent rate of children/month. 
2. Protocol delivery: The percent of on/off protocol children.  
3. Outcome completion: The percent of children with complete outcome measures. 
 
To evaluate acceptibilty, we will conduct end-of-study interviews with caregivers to assess outcomes 
related to the ease and utility of participating in the study, including our methods of data collection.  
 
Secondary outcomes: 
 
Our secondary aim is to explore patterns of recovery in children with SNI during the peri-operative 
period. We will assess: 
 
Objective 1: Function in children with SNI, including exploring patterns in specific domains of gain/loss. 
 
Objective 2: Quality of life in children with SNI and their families 
 
Objective 3: Profile of inflammatory biomarkers to compare children's inflammation levels peri-
operatively. 

 
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HYPOTHESIS 
We expect to confirm feasibility and acceptability of conducting peri-operative research in this 
population, with at least 12 children consented in 1 year and >85% protocol delivery and outcome 
completion. We anticipate to be able to measure PoFD, quality of life for children and caregivers, and 
inflammatory biomarkers.  
 
METHODOLOGY 
 
Study Design 
This study will use an observational, pre-post intervention design to evaluate our methods of 
documenting the post-operative functional trajectory and quality of life in children with SNI and their 
families during the peri-operative period. We will explore the functional abilities and quality of life of 
children with SNI and their parent/caregivers pre- and post- major orthopaedic surgery, and we will 
undertake measurements of inflammatory biomarkers 
 
Study Population 
Our study population will include 12 children with SNI and their parent/caregiver planning to undergo 
major orthopaedic surgery at the BC Children’s Hospital Orthopaedic Clinics. One parent/caregiver may 
participate per child participant (the same parent/caregiver throughout the study).  
 
Our study defines children with SNI as those with significant neurological disorders and evidence of 
developmental delay across all domains of daily function, who rely on assistive devices/technology for 
daily activities and well-being, such as mobility aids or feeding devices8 To accurately assess the 
functional status of our participants for screening pre-operatively, we will use the Gross Motor Function 
Classification System (GMFCS)30 and the Communication Function Classification System (CFCS)31 during 
participant screening. 
 
The GMFCS is a valid and reliable five-level standardized tool used to classify children and youth with SNI 
based on their level of movement ability30,32–34. For this study, we will include children classified at Levels 
II  to V. Children in Level II may have difficulty balancing on uneven terrain, inclines, in crowded or 
confined areas and may require physical assistance or mobility device for walking over long distances. 
Children in Level III will walk with hand-held mobility devices and use wheeled mobility when traveling 
long distances. Children in Level IV have severe limitations in their walking ability, including with 
assistance, and rely on a wheelchair most of the time. Children in Level V have profound physical 
impairments that impact their head and trunk postural control; they are unable to sit, stand, or walk 
independently and voluntarily.  
 
The CFCS is a valid and reliable five-level tool used to classify how individuals communicate with familiar 
and unfamiliar partners31. For our study, we will include children at Levels II   to V. Children at level II 
alternates between being sender and receiver but may converse slower. Children at level III will typically 
communicate effectively with familiar communication partners, but not with unfamiliar partners. 
Children at Level IV are inconsistent in their ability to communicate with familiar communication 
partners. Children at Level V are rarely able to communicate effectively, including with familiar people, 
and often require alternative communication methods to interact. 
 
“Major” orthopedic procedures are defined, for the purposes of this study, as operations that are equal 
to or greater than 90 minutes in “skin-to-skin" time (initial incision to skin closure). Orthopedic 
procedures (hip reconstruction and spine stabilization) are the most common major procedures that 

 
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children with SNI are exposed to, although they also experience many “minor” procedures, such as 
gastrostomy tube placement, tendon releases, and dental procedures. We are focusing on the longer 
procedures for this pilot study both because anecdotally, parents have not described the same degree of 
concern with shorter operations, and for methodological reasons, to improve the opportunity to identify 
this  this phenomenon.  
   
Study participant eligibility criteria 
 
Inclusion Criteria 
1) Children with SNI, including the Gross Motor Function Classification System (GMFCS) levels II to 
V and Communication Function Classification System (CFCS) levels II to V, and; 
2) undergoing major orthopaedic surgical management of musculoskeletal pathology (spine or hip; 
major defined as a greater than or equal to 90 minute procedure with skin-to-skin contact) at 
the BC Children's Hospital and; 
3) between 5 and 18 years at the time of their surgical procedure  
 
Exclusion Criteria 
1) GMFCS level I and CFCS level I 
2) Non-English-speaking Parents/Caregivers 
 
Control group - justifications and eligibility criteria 
We aim to recruit 5 control participants specifically for our biomarker analysis (ie. we will only collect 
demographics and blood samples). The inclusion of this control group allows for a direct comparison of 
the pre-, peri-, and post-operative inflammatory responses between children with SNI vs neurotypical 
patients undergoing similar procedures. This is essential for discerning whether any observed altered 
biomarkers levels specific to children with SNI (ie. and not seen in neurotypical patients), or if they 
reflect a generalized postoperative inflammatory response. This comparison strengthens the 
interpretation of our biomarker findings by providing a baseline, which will reduce the risk of 
misattributing inflammation-related findings solely to the presence of SNI rather than the general 
surgical response. Additionally, the inclusion of controls improves the clinical relevance of our study, as 
it will help determine whether children with SNI exhibit exaggerated, prolonged, or distinct 
inflammatory responses compared to their neurotypical counterparts. These insights will be critical for 
guiding future research and potential interventions aimed at mitigating PoFD in children with SNI.  
 
Inclusion Criteria 
1) Neurotypically developing children (GMFCS I and CFCS I) 
2) Undergoing major orthopaedic surgical management of musculoskeletal pathology (spine or hip; 
major defined as a greater than or equal to 90 minute procedure with skin-to-skin contact) at 
the BC Children's Hospital 
3) Between 5 and 18 years of age at the time of their surgical procedure  
 
Exclusion Criteria 
1) Non-English-speaking parents/caregivers 
 
Participant Screening and Recruitment 
Participants for this study will be recruited in-person through the BC Children’s Hospital Orthopaedic 
Clinics, including the Orthopedic Cerebral Palsy Clinic (CP Clinic) and the Spine Clinic. Study participants 

 
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will receive $100 remuneration for their participation in the study, and control participants will receive 
$50 remuneration for their participation. 
 
Participant Screening 
 
1) Screening through standard-of-care clinical visits  
 
The BCCH Orthopedic Clinic (Hip and Spine) will serve as the main recruitment site, where children with 
SNI and neurotypical children with scoliosis attend regular orthopedic visits. Prospective participants will 
be identified from a list of patients who will be attending a regular clinic visit at both the Orthopedic 
Cerebral Palsy Clinic and Spine Program by respective clinic Research Assistants (RAs). Following 
identification, the RAs at each respective clinic will screen patients as standard for their own studies and 
flag patients that align with our study’s inclusion criteria; for the Hip Clinic, this will be for the Hippy 
Lab’s Outcomes of Hip Interventions for Children with Cerebral Palsy – An International Multi-Centre 
Prospective Comparative Cohort Study (Ethics ID # H14-01402) study using PowerChart prior to patients’ 
clinical visits, and Spine Program RAs will screen patients independently. The Clinic RAs will then provide 
the Siden Lab RA with a list of their potential participants with information relevant to the Post-Op 
Changes Study. Using this list, the Siden Lab RA will then screen for Post-Op Changes Study inclusion 
criteria (age, GMFCS and CFCS status, on track for major hip/spine surgery), and will coordinate with 
Clinic RAs to visit these participants at their upcoming pre-operative appointment. 
 
2) Screening through surgical slate – Hip Clinic 
 
The Siden Lab RA will receive an operating room slate for the patients scheduled for surgery in the 
upcoming month from the Hip and Spine Clinics’ Medical Office Assistant(s), along with the dates of 
surgery and pre-operative appointments. These participants may then be screened for our inclusion 
criteria by the Siden Lab Research Nurse using PowerChart; for eligible participants, arrangements will 
then be made for participants to be approached and recruited at their pre-operative appointment. 
 
3) Screening through CST Cerner 
 
The Medical Student Research Assistant with Level 2 Cerner access will filter their search on CST Cerner 
specifically for the 3 surgeons whose patients we have obtained permission to recruit from: Drs. Andrea 
Simmonds, Firoz Miyanji, and Kishore Mulpuri. This will result in a list of the operating schedule and pre-
operative appointments for each surgeon. The RA will then access the patients' charts for those who 
have a confirmed surgery date or pre-operative visit to screen for our inclusion criteria to determine if 
they are eligible to be a study participant or a control. Screened eligible patients will be confirmed with 
the Orthopaedic team RAs prior to approaching the patient. With this, we will track all patients who 
would be eligible, as well as patients that we approached, in a password protected Excel Spreadsheet 
housed under our lab's V Drive. We have attached the spreadsheet under Recruitment documents 
which includes all data we will be collecting as part of this screening. 
 
Participant Recruitment 
 
1) Recruitment at regular clinic visit: 
 
During the standard-of-care clinical visit of patients who will likely undergo surgery within ~6 months (as 
identified by the RAs/team of each clinic conducting screening), the patient’s clinician will ask the 

 
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patient’s parent/caregiver if they are interested in participating in research and will briefly introduce 
them to the study. If the patient’s parent/caregiver is interested, the RA will approach the patient and 
their parent/caregiver regarding the details of the study. If the parent/caregiver is interested in 
participating, the RA will then obtain consent from the participants (the parent/caregiver may provide 
consent on behalf of their child) either electronically on a tablet as an E-Consent through REDCap or on 
paper. If this recruitment visit falls within three months of the study participant’s scheduled surgery, the 
RA may then proceed to take baseline/outcome measurements. If the recruitment visit falls outside 
three months before the patient’s surgery date, the RA may obtain demographic information, and 
baseline outcome measurements will be obtained at another clinical or pre-operative visit within three 
months of surgery (for study participants). The patient’s parent/caregiver may alternatively request 
additional time to consider participating; in this case, they will be provided with a paper copy of the 
study details and consent form, and may be approached at their next clinical visit to re-determine 
interest. If they are controls and data collection can occur at the same time of recruitment and consent, 
then only demographic information will be collected. 
 
2) Recruitment at pre-operative clinic visit: 
 
After confirming patient’s eligibility, the RA may approach the patient and their parent/caregiver 
regarding study participation during their pre-operative visit. During their pre-operative clinical visit, the 
patient’s clinician will ask the patient’s parent/caregiver if they are interested in participating in research 
and will briefly introduce them to the study. If the patient’s parent/caregiver is interested, the RA will 
approach the patient and their parent/caregiver regarding the details of the study. If the 
parent/caregiver is interested in participating, the RA will then obtain consent from the participants (the 
parent/caregiver may provide consent on behalf of their child) either electronically on a tablet as an E-
Consent through REDCap or on paper. The RA may then proceed to take baseline and outcome 
measurements for study participants, and only demographic information for control participants. The 
patient’s parent/caregiver may alternatively request additional time to consider participating; in this 
case, they will be emailed a REDCap link to the consent form that they can fill out from home 
electronically. Once they have been consented, they can then receive a link to our relevant 
questionnaires (to be filled before surgery). The RA will then coordinate with the patient’s MOA to 
ensure that the study visit timeline are timed congruently with clinical post-operative follow-up visits. 
 
3) Recruitment on day of surgery 
 
After confirming the patient’s eligibility, if it is not possible to recruit at the pre-operative appointment 
the RA may approach the patient and their parent/caregiver regarding study participation on the day of 
their surgery prior to when the surgery begins. If the parent/caregiver and child agrees to participate, 
the RAs may obtain consent/assent and complete the baseline/outcome measurements at this time. The 
RA will then monitor CST Cerner to ensure that the clinical post-operative follow-up visits are timed 
congruently with the study visit timeline. 
 
Recruitment video 
At all three recruitment types, we will show our recruitment video to the study participants (children 
with SNI) to provide them with more information about the study prior to their review/signing of the 
consent form. 
 
Sample Size Justification 

 
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We aim to recruit 12 study participants and 5 control participants for this pilot study, which will be 
useful to 1) test the feasibility and acceptibility of our methods and tools for later use on a larger scale, 
and 2) allow us to describe patterns in preliminary data to identify early associations that may guide the 
methodology of a larger study. A sample size of 12 study participants and 5 control participants is likely 
feasible, as the Orthopedic Clinic sees an average of 50 children for hip/spine surgery each year. 
 
STUDY PROCEDURES 
The study will include a total of six timed assessments (Figure 1). This includes three primary study 
assessments to collect functional survey and biomarker data: 1) at baseline (within three months of 
surgery), 2) 6 weeks post-operatively, and 3) 6 months post-operatively. Additionally, we will conduct 
three biomarker-specific collections during the time of surgery immediately prior to the initiation of the 
procedure, 12-24 hours post-operatively, and 1-2 weeks post-operatively, based on the physiology of 
the sequence of the innate and adaptive immune response systems. Most study assessments will be 
timed at standard of care visits at the BC Children’s Hospital. Each primary study visit will take 
approximately 25-30 minutes. Total time of participation will be 1.5 – 2 hours. Measures will be taken 
congruently with standard clinical care data collection where possible to reduce participation burden. 
Some participants may be recruited exclusively for functional/quality of life data collection while the 
team awaits approvals for blood collection and storage, to allow for an increased sample size. 
 
Baseline Data Collection 
Study participants who have consented will undergo a total of 3 primary study visits and 6 biomarker 
collections (Figure 1). At Study assessment #1, demographic information for the child (including 
variables such as age, sex, race/ethnicity, primary diagnosis, GMFCS/CFCS/MACS score) and 
demographic information of the parent/caregiver (age, sex, relationship to child, employment status, 
household income, race/ethnicity, distance from home to primary medical facility) will be collected. 
Outcome measures (see below) will also be collected. Baseline data collection aims to capture the initial 
status of participants before any surgical intervention, providing a reference point for post-operative 
comparisons.  
 
Outcome Measures 
The follow study procedures correspond to our primary aims: 
 
To assess feasibility, we will evaluate three primary feasibility outcomes at the time of data analysis. We 
will calculate consent rate, which is the overall average consent rate of children/month; protocol 
delivery, which is the percent of on/off protocol children; and outcome completion, the percent of 
children with complete outcome measures. 
 
To assess acceptability, anytime after the 6-week timepoint , we will conduct semi-structured interviews 
with families to assess their experience in participating in this study. We will also ask families if there 
were any domains that were not included in our standardized functional/quality of life questionnaire 
and questions. Parents may select from pre-identified domains as prompts (including pain, energy, and 
sleep), and may also provide additional insight through open-ended conversation. Interviews will be 
recorded, transcribed, and analyzed using thematic analysis.  
 
The follow study procedures correspond to our secondary aims: 
 
Our primary tool for this study to measure our outcomes of function and quality of life will be the 
Caregivers Priorities and Child Health Index of Life with Disabilities (CPCHILD) Questionnaire35, which has 

 
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been proved to be both valid and reliable35,36. The CPCHILD Questionnaire is used to assess the 
effectiveness of interventions aimed at improving and preserving outcomes for children aged 5-18 years 
old with severe disabilities, based on caregivers’ perspectives. This questionnaire includes 37 items 
collected from the parent/caregiver measured across six domains: Personal Care/Activities of Daily 
Living; Positioning, Transferring, and Mobility; Comfort and Emotions; Communication and Social 
Interaction; Health; Child’s Overall Quality of Life; with additional sections for Importance of Items to 
Child’s Quality of Life, Facts about the Child, and Facts about the Caregiver. From responses, 
standardized scores can be retrieved on a scale of 0 to 100 for each domain, as well as a total score, and 
we will be examining comparisons at the item-level pre/post-operatively.  
 
Parents/caregivers will also be asked to answer two supplemental questions to provide further insight 
into 1) other areas of function that were not described by CPCHILD and 2) the quality of life of 
caregivers. 
 
Blood collection of study participants 
 
To collect information regarding the peri-operative inflammatory profile, we will collect blood samples 
from study participants at all study time points: 
 
• 
Timepoint #1 (pre-operative appointment): during routine care through venipuncture by C&W 
Lab Technicians. An additional finger prick will be taken by technicians (non-routine).  
• 
Timepoint #2 (surgery): prior to incision through peripheral venous line by OR staff 
• 
Timepoint #3 (12-24 hours post-op): by bedside nurses in T6 or PICU 
• 
Timepoint #4 (1-2 weeks post-op) 
o Hip patients: non-routine collection by C&W Lab  
o Spine patients: by bedside nurses in T6 
• 
Timepoint #5 (6 weeks post-op): non-routine by C&W Lab  
• 
Timepoint #6 (6 months post-op): non-routine by C&W Lab 
 
Participants may choose not to provide their blood samples and remain a participant for our functional 
questionnaire and quality of life-related data collection. Analysis of blood will be done in collaboration 
with the Core Facility for Neurology Biomarker Innovation/Wellington Laboratory. 
 
A 3 mL aliquot of the child’s blood will be set aside for our study. This blood will be transferred to the 
appropriate collection tubes (heparin coated) to prevent clotting and will be transported to  the BC 
Children’s Hospital BioBank for storage. 
 
A lancing device will be used for finger capillary sampling to collect 50uL of blood from the child. The 
blood sample will be immediately collected following puncture using capillary tubes and transferred into 
pre-labeled collection tubes (heparin coated to prevent clotting) and mixed to ensure the sample 
remains homogeneous. After collection, the sample will be  stored at –80 C at the BC Children’s Hospital 
BioBank. 
 
Blood collection of control participants 
Control participants will only have their blood drawn at three timpoints. Relative to study participants, 
blood samples will be collected from controls at: 
 

 
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• 
Timepoint #2 (surgery): prior to incision through peripheral venous line by OR staff 
• 
Timepoint #3 (12-24 hours post-op): through routine venipuncture in T6 
• 
Timepoint #6 (6 months post-op): non routine bloodwork by C&W lab  
 
At the conclusion of data collection, all blood samples will be transported to the UBC Wellington 
Laboratory (coordinated by the BC Children’s Hospital BioBank). 
 
Further, our Research Nurse will extract medication, surgery, and anesthetic-specific data (e.g. pain 
management drugs [anti-inflammatory, steroids, and immune-suppresants], surgery duration, 
anaesthesia-specific exposure) pre- and post-operatively from participants’ electronic medical records 
for our team to better contextualize PoFD and inflammatory response. 
 
Timeline 
Measures will be taken at baseline (within three months of surgery), at the time of operation, 12-24 
hours following operation, 1-2 weeks following operation, 6 weeks post-operatively and 6 months post-
operatively as per the schedule below. Study visits will be timed congruently with standard clinical care 
visits or pre-operative/post-operative clinic visits. 
 
 
Figure 1. Study timeline for study participants 
 
DATA ANALYSIS 
 
Data analysis will focus primarily on evaluating the feasibility and acceptability of the study protocol. 
Baseline descriptive data will be presented as median and interquartile ranges, and categorical data by 
percentages. Primary feasibility outcomes (consent rate, protocol delivery, and outcome completion 
rates) will be described as percentages. Acceptability data will be analyzed from semi-structured 
interviews. Qualitative data from interviews with families will be analyzed using thematic analysis to 
identify common themes and insights regarding their experiences with the study protocol.  
  
For secondary outcomes related to improvement and/or decline, we will describe patterns in 1) child’s 
function, 2) quality of life of child and caregiver, and 3) inflammatory profile pre- and post-operatively. 
Given our small sample size, we cannot statistically confirm correlations of PoFD and inflammatory 
biomarkers, though we may detect patterns in specific markers. 
 
In our analysis (all outcome measures), we hope to stratify our results by sub-category of GMFCS 5, as 
has been done previously. The sub-categories are presence of a gastrostomy tube, a tracheostomy, 

 
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history of seizures, and nonverbal status. Patients with 0 impairments will be classified as GMFCS level 
5.0, patients with 1 impairment as level 5.1, patient with 2 impairments as level 5.2, and patients with 3 
or more impairments as level 5.3. This stratification will allow us to examine participants’ response to 
surgery based on an indicator of severity of their neurological impairment.  
 
Blood collection 
 
In collaboration with facilities and technologies of the UBC Wellington Lab, biospecimens will be 
analyzed on the Alamar Biosciences ARGO HT platform, a fully automated immunoassay platform 
designed for NUcleic acid Linked Immuno-Sandwich Assay (NULISA) technology. The NULISA technology 
is a ligation-based assay which suppresses assay background using a dual capture and release 
mechanism with oligonucleotide-conjugated antibodies. It is designed to detect proteins across a high 
dynamic range with high sensitivity through next-generation sequencing of barcoded reporter DNA 
produced by the assay which also allows high multiplex capability. Analysis will include relative 
quantitation of a panel of 120 analytes designed to profile major hallmarks of central nervous system 
diseases (CNS) using the NULISAseq CNS Panel 120 assay. This assay is commercially available from 
Alamar Biosciences, and has been validated for sensitivity, precision, cross-lot, and cross-machine 
performance. Validation testing includes dynamic range, specificity, spike recovery, dilution linearity, 
sensitivity, intra-run, inter-plate, and inter-lot precision and accuracy. Each NULISA assay run contains 4 
negative controls (NCs), 3 sample controls, 3 inter-plate controls (IPCs), and up to 86 samples. Each 
sample is spiked with the same concentration of an internal control (IC) used in the normalization 
procedure and as a method for assessing the uniformity of the assay run. IPC samples are pooled plasma 
controls used to normalize samples between experiments and to assess performance of the assay. NC 
wells are reactions where no sample input is provided and is used to assess limit of detection (LOD) for 
each target assay and to assess run quality. SC samples are pooled plasma controls from an independent 
source different from IPCs. SCs are used for calculating intra- and inter-plate coefficient of variation (CV). 
Specimens are assayed in singlicate in randomized order with no dilution required. Quality control 
measures, including but not limited to, CV of spiked IC signal across all samples, CV of the total assay 
signal across all IPCs, Median CV of all IPC targets, and percentage of targets that are detectable, defined 
as detectable if > 50% samples have signal > limit of detection, are recorded every run. Raw sequencing 
reads are processed to generate target- and sample-specific counts called NULISA Protein Quantification 
(NPQ) values. The raw sequencing reads are first normalized to the IC to control for well-to-well 
variation and then normalized to the median IC-noramlized counts for the IPC samples for plate-to-plate 
variation.  
 
With respect to statistical analysis of our control group, our primary analysis will involve a group 
comparison at each timepoint to assess whether, and to what extent, any biomarkers are altered 
(increased or decreased) in children with SNI compared to our controls. Given the large biomarker panel 
(approximately 120 biomarkers), data visualization using volcano plots will be employed. Multiple 
comparison adjustments will be applied using the false discovery rate (FDR) correction method 
(Benjamini-Hochberg). Biomarker significance will be determined based on effect size thresholds, 
specifically log2(fold change), with a significance level set at 5% (Type I error). Additionally, associations 
between biomarkers and study variables (e.g., age, sex) will be explored using Wilcoxon rank-sum tests. 
 
Outcomes 
The following outcomes will be utilized to deem our study feasible and acceptable: 
 
Feasibility: 

 
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1. Consent rate: An average consent rate of 2-3 children/month 
2. Protocol delivery: The protocol can be implemented with >85% protocol delivery. 
3. Outcome completion: >85% of outcomes will be complete and measured as scheduled. 
 
Acceptability: 
4. General positive report on participation based on semi-structured interview 
 
Secondary outcomes; we will explore pre- and post-operative data to describe patterns in: 
1. Function of the child, as scored on the CPCHILD; 
2. Quality of life of caregiver, as described by supplemental question; 
3. Inflammatory profile of child 
 
DATA STORAGE 
 
All participants will receive unique study ID numbers to be used on all written forms and samples to 
maintain confidentiality. A master list linking identifiers to study codes will be kept separately in a 
password protected electronic location with the Siden Lab Research Coordinator. All de-identified data 
will be transcribed and stored in a secure, password-protected REDCap database with access limited to 
the authorized study team. Any hard copy data will be stored in locked filing cabinets with the Siden Lab 
at BC Children’s Hospital. Hard copy documents and data will be kept for five years following study 
completion and then destroyed. Biospecimen will be securely stored until they are destroyed at the UBC 
Wellington Lab. Informed consent documents will describe the measures taken to protect participant 
data, and participants will be assured that their information will be used solely for the purposes of this 
study. All electronic communications involving participant data will use encrypted channels to prevent 
unauthorized access. 
 
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